U.S. patent number 8,440,820 [Application Number 13/348,500] was granted by the patent office on 2013-05-14 for antioxidant inflammation modulators: oleanolic acid derivatives with saturation in the c-ring.
This patent grant is currently assigned to Reata Pharmaceuticals, Inc.. The grantee listed for this patent is Eric Anderson, Xin Jiang, Xiaofeng Liu, Melean Visnick. Invention is credited to Eric Anderson, Xin Jiang, Xiaofeng Liu, Melean Visnick.
United States Patent |
8,440,820 |
Anderson , et al. |
May 14, 2013 |
**Please see images for:
( Certificate of Correction ) ** |
Antioxidant inflammation modulators: oleanolic acid derivatives
with saturation in the C-ring
Abstract
This invention provides, but is not limited to, novel oleanolic
acid derivatives having the formula: ##STR00001## wherein the
variables are defined herein. Also provided are pharmaceutical
compositions, kits and articles of manufacture comprising such
compounds, methods and intermediates useful for making the
compounds, and methods of using the compounds and compositions.
Inventors: |
Anderson; Eric (Southlake,
TX), Jiang; Xin (Dallas, TX), Liu; Xiaofeng (Dallas,
TX), Visnick; Melean (Irving, TX) |
Applicant: |
Name |
City |
State |
Country |
Type |
Anderson; Eric
Jiang; Xin
Liu; Xiaofeng
Visnick; Melean |
Southlake
Dallas
Dallas
Irving |
TX
TX
TX
TX |
US
US
US
US |
|
|
Assignee: |
Reata Pharmaceuticals, Inc.
(Irving, TX)
|
Family
ID: |
41726396 |
Appl.
No.: |
13/348,500 |
Filed: |
January 11, 2012 |
Prior Publication Data
|
|
|
|
Document
Identifier |
Publication Date |
|
US 20120245374 A1 |
Sep 27, 2012 |
|
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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13033452 |
Feb 23, 2011 |
8124656 |
|
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12426737 |
Apr 20, 2009 |
7915402 |
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61046332 |
Apr 18, 2008 |
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61111333 |
Nov 4, 2008 |
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Current U.S.
Class: |
540/519; 548/143;
568/368; 562/498; 548/136; 560/116; 564/188; 564/148; 540/520;
549/354; 548/250 |
Current CPC
Class: |
A61P
13/12 (20180101); A61P 9/10 (20180101); C07J
63/008 (20130101); C07C 69/757 (20130101); A61P
3/00 (20180101); A61P 3/10 (20180101); A61P
31/00 (20180101); A61P 17/06 (20180101); A61P
9/12 (20180101); C07D 413/08 (20130101); A61P
19/02 (20180101); A61P 25/00 (20180101); A61P
25/28 (20180101); A61P 9/00 (20180101); A61P
29/00 (20180101); C07D 261/20 (20130101); A61P
1/00 (20180101); A61P 35/02 (20180101); C07D
207/27 (20130101); A61P 3/04 (20180101); A61P
25/16 (20180101); A61P 35/00 (20180101) |
Current International
Class: |
C07C
49/115 (20060101); C07D 313/06 (20060101); C07D
271/10 (20060101); C07D 285/12 (20060101); C07C
69/753 (20060101); C07C 61/125 (20060101); C07D
491/22 (20060101); C07D 257/04 (20060101); C07C
233/00 (20060101) |
Field of
Search: |
;540/519,520
;548/136,143,250 ;549/354 ;560/116 ;562/498 ;564/148,188 ;558/429
;568/368 |
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Aug 2010 |
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WO |
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|
Primary Examiner: Shterengarts; Samantha
Attorney, Agent or Firm: Parker Highlander PLLC
Parent Case Text
The present application is a continuation application of co-pending
application Ser. No. 13/033,452, filed Feb. 23, 2011, which is a
continuation of application Ser. No. 12/426,737, filed Apr. 20,
2009, now U.S. Pat. No. 7,915,402, which claims the priority of
U.S. Provisional Application Nos. 61/046,332, filed Apr. 18, 2008,
and 61/111,333, filed Nov. 4, 2008, the entire contents of each of
which are incorporated herein by reference.
Claims
What is claimed is:
1. A compound of the formula: ##STR00088## wherein: Y is cyano or
--C(O)R.sub.a, further wherein: R.sub.a is: hydrogen, hydroxy,
halo, amino, azido, mercapto or silyl; or alkyl.sub.(C.ltoreq.12),
alkenyl.sub.(C.ltoreq.12), alkynyl.sub.(C.ltoreq.12),
aryl.sub.(C.ltoreq.12), aralkyl.sub.(C.ltoreq.12),
heteroaryl.sub.(C.ltoreq.12), heteroaralkyl.sub.(C.ltoreq.12),
alkoxy.sub.(C.ltoreq.12), alkenyloxy.sub.(C.ltoreq.12),
alkynyloxy.sub.(C.ltoreq.12), aryloxy.sub.(C.ltoreq.12),
aralkoxy.sub.(C.ltoreq.12), heteroaryloxy.sub.(C.ltoreq.12),
heteroaralkoxy.sub.(C.ltoreq.12), acyloxy.sub.(C.ltoreq.12),
alkylamino.sub.(C.ltoreq.12), alkoxyamino.sub.(C.ltoreq.12),
dialkylamino.sub.(C.ltoreq.12), alkenylamino.sub.(C.ltoreq.12),
alkynylamino.sub.(C.ltoreq.12), arylamino.sub.(C.ltoreq.12),
aralkylamino.sub.(C.ltoreq.12), heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12), or a
substituted version of any of these groups; X is OR.sub.b,
NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and R.sub.c are each
independently: hydrogen or hydroxy; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.b is absent when the atom to which it is bound is part
of a double bond, further provided that when R.sub.b is absent the
atom to which it is bound is part of a double bond; R.sub.1 is:
hydrogen, cyano, hydroxy, halo or amino; or alkyl.sub.(C.ltoreq.8),
alkenyl.sub.(C.ltoreq.8), alkynyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), heteroaralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and R.sub.4 is
alkyl.sub.(C.ltoreq.8) or substituted alkyl.sub.(C.ltoreq.8); or a
pharmaceutically acceptable salt thereof.
2. The compound of claim 1, further defined as: ##STR00089##
wherein: R.sub.a is: hydrogen, hydroxy, halo, or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
alkenyloxy.sub.(C.ltoreq.8), alkynyloxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), aralkoxy.sub.(C.ltoreq.8),
heteroaryloxy.sub.(C.ltoreq.8), heteroaralkoxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
alkoxyamino.sub.(C.ltoreq.8), dialkylamino.sub.(C.ltoreq.8),
alkenylamino.sub.(C.ltoreq.8), alkynylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), aralkylamino.sub.(C.ltoreq.8),
heteroarylamino.sub.(C.ltoreq.8),
heteroaralkylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; X is OR.sub.b or
NR.sub.bR.sub.c, wherein R.sub.b and R.sub.c are each
independently: hydrogen or hydroxy; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.b is absent when the atom to which it is bound is part
of a double bond, further provided that when R.sub.b is absent the
atom to which it is bound is part of a double bond; and R.sub.2 is:
cyano, hydroxy, halo or amino; or fluoroalkyl.sub.(C.ltoreq.8),
alkenyl.sub.(C.ltoreq.8), alkynyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a pharmaceutically acceptable salt thereof.
3. The compound of claim 2, further defined as: ##STR00090##
wherein: R.sub.a is: hydrogen, hydroxy, halo, or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
alkenyloxy.sub.(C.ltoreq.8), alkynyloxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), aralkoxy.sub.(C.ltoreq.8),
heteroaryloxy.sub.(C.ltoreq.8), heteroaralkoxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
alkoxyamino.sub.(C.ltoreq.8), dialkylamino.sub.(C.ltoreq.8),
alkenylamino.sub.(C.ltoreq.8), alkynylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), aralkylamino.sub.(C.ltoreq.8),
heteroarylamino.sub.(C.ltoreq.8),
heteroaralkylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and R.sub.2 is: cyano
or fluoro; or fluoroalkyl.sub.(C.ltoreq.5),
alkenyl.sub.(C.ltoreq.5), alkynyl.sub.(C.ltoreq.5),
heteroaryl.sub.(C.ltoreq.5), acyl.sub.(C.ltoreq.5),
acyloxy.sub.(C.ltoreq.5), amido.sub.(C.ltoreq.5), or a substituted
version of any of these groups; or a pharmaceutically acceptable
salt thereof.
Description
BACKGROUND OF THE INVENTION
I. Field of the Invention
The present invention relates generally to the fields of biology
and medicine. More particularly, it concerns compounds and methods
for the treatment and prevention of diseases such as those
associated with oxidative stress and inflammation.
II. Description of Related Art
Many serious and intractable human diseases are associated with
dysregulation of inflammatory processes, including diseases such as
cancer, atherosclerosis, and diabetes, which were not traditionally
viewed as inflammatory conditions. Similarly, autoimmune diseases
such as rheumatoid arthritis, lupus, psoriasis, and multiple
sclerosis involve inappropriate and chronic activation of
inflammatory processes in affected tissues, arising from
dysfunction of self vs. non-self recognition and response
mechanisms in the immune system. In neurodegenerative diseases such
as Alzheimer's and Parkinson's diseases, neural damage is
correlated with activation of microglia and elevated levels of
pro-inflammatory proteins such as inducible nitric oxide synthase
(iNOS).
One aspect of inflammation is the production of inflammatory
prostaglandins such as prostaglandin E, whose precursors are
produced by the enzyme cyclo-oxygenase (COX-2). High levels of
COX-2 are found in inflamed tissues. Consequently, inhibition of
COX-2 is known to reduce many symptoms of inflammation and a number
of important anti-inflammatory drugs (e.g., ibuprofen and
celecoxib) act by inhibiting COX-2 activity. Recent research,
however, has demonstrated that a class of cyclopentenone
prostaglandins (e.g., 15-deoxy prostaglandin J2, a.k.a. PGJ2) plays
a role in stimulating the orchestrated resolution of inflammation.
COX-2 is also associated with the production of cyclopentenone
prostaglandins. Consequently, inhibition of COX-2 may interfere
with the full resolution of inflammation, potentially promoting the
persistence of activated immune cells in tissues and leading to
chronic, "smoldering" inflammation. This effect may be responsible
for the increased incidence of cardiovascular disease in patients
using selective COX-2 inhibitors for long periods of time.
Corticosteroids, another important class of anti-inflammatory
drugs, have many undesirable side effects and frequently are not
suitable for chronic use. Newer protein-based drugs, such as
anti-TNF monoclonal antibodies, have proven to be effective for the
treatment of certain autoimmune diseases such as rheumatoid
arthritis. However, these compounds must be administered by
injection, are not effective in all patients, and may have severe
side effects. In many severe forms of inflammation (e.g., sepsis,
acute pancreatitis), existing drugs are ineffective. In addition,
currently available drugs typically do not have significant
antioxidant properties, and are not effective in reducing oxidative
stress associated with excessive production of reactive oxygen
species and related molecules such as peroxynitrite. Accordingly,
there is a pressing need for improved therapeutics with antioxidant
and anti-inflammatory properties.
A series of synthetic triterpenoid analogs of oleanolic acid have
been shown to be inhibitors of cellular inflammatory processes,
such as the induction by IFN-.gamma. of inducible nitric oxide
synthase (iNOS) and of COX-2 in mouse macrophages. See Honda et al.
(2000a); Honda et al. (2000b), and Honda et al. (2002), which are
all incorporated herein by reference. For example, one of these,
2-cyano-3,12-dioxooleane-1,9(11)-dien-28-oic acid methyl ester
(CDDO-Me), is currently in clinical trials for a variety of
disorders related to inflammation, including cancer and diabetic
nephropathy. The pharmacology of these molecules is complex, as
they have been shown to affect the function of multiple protein
targets and thereby modulate the function of several important
cellular signaling pathways related to oxidative stress, cell cycle
control, and inflammation (e.g., Dinkova-Kostova et al., Ahmad et
al., 2006; Ahmad et al., 2008; Liby et al.,). Given that the
biological activity profiles of the known oleanolic acid
derivatives vary, and in view of the wide variety of diseases that
may be treated with compounds having potent antioxidant and
anti-inflammatory effects, it is desirable to synthesize new
candidates for the treatment or prevention of disease.
SUMMARY OF THE INVENTION
The present disclosure provides new compounds with antioxidant and
anti-inflammatory properties, methods for their manufacture, and
methods for their use. Compounds covered by the generic or specific
formulas below or specifically named are referred to as "compounds
of the invention," "compounds of the present disclosure," "the
present oleanolic acid derivatives" in the present application.
In some aspects, the disclosure provides compounds of the
formula:
##STR00002## wherein: Y is cyano, heteroaryl.sub.(C.ltoreq.12),
substituted heteroaryl.sub.(C.ltoreq.12), or --C(O)R.sub.a, further
wherein R.sub.a is: hydrogen, hydroxy, halo, amino, hydroxyamino,
azido, silyl or mercapto; alkyl.sub.(C.ltoreq.12),
alkenyl.sub.(C.ltoreq.12), alkynyl.sub.(C.ltoreq.12),
aryl.sub.(C.ltoreq.12), aralkyl.sub.(C.ltoreq.12),
heteroaryl.sub.(C.ltoreq.12), heteroaralkyl.sub.(C.ltoreq.12),
alkoxy.sub.(C.ltoreq.12), alkenyloxy.sub.(C.ltoreq.12),
alkynyloxy.sub.(C.ltoreq.12), aryloxy.sub.(C.ltoreq.12),
aralkoxy.sub.(C.ltoreq.12), heteroaryloxy.sub.(C.ltoreq.12),
heteroaralkoxy.sub.(C.ltoreq.12), acyloxy.sub.(C.ltoreq.12),
alkylamino.sub.(C.ltoreq.12), dialkylamino.sub.(C.ltoreq.12),
alkoxyamino.sub.(C.ltoreq.12), alkenylamino.sub.(C.ltoreq.12),
alkynylamino.sub.(C.ltoreq.12), arylamino.sub.(C.ltoreq.12),
aralkylamino.sub.(C.ltoreq.12), heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12),
alkylsulfonylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12),
alkylthio.sub.(C.ltoreq.12), alkenylthio.sub.(C.ltoreq.12),
alkynylthio.sub.(C.ltoreq.12), arylthio.sub.(C.ltoreq.12),
aralkylthio.sub.(C.ltoreq.12), heteroarylthio.sub.(C.ltoreq.12),
heteroaralkylthio.sub.(C.ltoreq.12), acylthio.sub.(C.ltoreq.12),
alkylammonium.sub.(C.ltoreq.12), alkylsulfonium.sub.(C.ltoreq.12),
alkylsilyl.sub.(C.ltoreq.12), or a substituted version of any of
these groups; or R.sub.a comprises a nitrogen atom that is also
attached to carbon atom 13 and R.sub.d to form:
##STR00003## wherein R.sub.d is alkyl.sub.(C.ltoreq.8),
alkenyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), or a substituted version of any of
these groups; Z is a single or double bond, --O-- or --NR.sub.e--,
wherein R.sub.e is hydrogen, hydroxy, alkyl.sub.(C.ltoreq.8) or
alkoxy.sub.(C.ltoreq.8); X is OR.sub.b, NR.sub.bR.sub.c, or
SR.sub.b, wherein R.sub.b and R.sub.c are each independently:
hydrogen or hydroxy; alkyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or a substituent
convertible in vivo to hydrogen; provided that R.sub.b is absent
when the atom to which it is bound is part of a double bond,
further provided that when R.sub.b is absent the atom to which it
is bound is part of a double bond; R.sub.1 is: hydrogen, cyano,
hydroxy, halo or amino; or alkyl.sub.(C.ltoreq.8),
alkenyl.sub.(C.ltoreq.8), alkynyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), heteroaralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; R.sub.3 is: absent or
hydrogen; alkyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8), or a substituted
version of any of these groups; or a substituent convertible in
vivo to hydrogen; provided that R.sub.3 is absent when the oxygen
atom to which it is bound is part of a double bond, further
provided that when R.sub.3 is absent the oxygen atom to which it is
bound is part of a double bond; R.sub.4 and R.sub.5 are each
independently alkyl.sub.(C.ltoreq.8) or substituted
alkyl.sub.(C.ltoreq.8); R.sub.6 is hydrogen, hydroxy or oxo;
R.sub.7 is hydrogen or hydroxy; and R.sub.8, R.sub.9, R.sub.10 and
R.sub.11 are each independently hydrogen, hydroxy,
alkyl.sub.(C.ltoreq.8), substituted alkyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8) or substituted alkoxy.sub.(C.ltoreq.8); or
pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00004## wherein:
Y is cyano, or --C(O)R.sub.a, further wherein: R.sub.a is:
hydrogen, hydroxy, halo, amino, hydroxyamino, azido or mercapto; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
dialkylamino.sub.(C.ltoreq.12), alkoxyamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12),
alkylsulfonylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12),
alkylthio.sub.(C.ltoreq.12), alkenylthio.sub.(C.ltoreq.12),
alkynylthio.sub.(C.ltoreq.12), arylthio.sub.(C.ltoreq.12),
aralkylthio.sub.(C.ltoreq.12), heteroarylthio.sub.(C.ltoreq.12),
heteroaralkylthio.sub.(C.ltoreq.12), acylthio.sub.(C.ltoreq.12),
alkylammonium.sub.(C.ltoreq.12), alkylsulfonium.sub.(C.ltoreq.12),
alkylsilyl.sub.(C.ltoreq.12), or a substituted version of any of
these groups;
X is OR.sub.b, NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and
R.sub.c are each independently: hydrogen or hydroxy;
alkyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or a substituent
convertible in vivo to hydrogen; provided that R.sub.b is absent
when the atom to which it is bound is part of a double bond,
further provided that when R.sub.b is absent the atom to which it
is bound is part of a double bond;
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.3 is: absent or hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.3 is absent when the oxygen atom to which it is bound is
part of a double bond, further provided that when R.sub.3 is absent
the oxygen atom to which it is bound is part of a double bond;
R.sub.4 and R.sub.5 are each independently alkyl.sub.(C.ltoreq.8)
or substituted alkyl.sub.(C.ltoreq.8); and
R.sub.6 and R.sub.7 are each independently hydrogen or hydroxy;
or pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00005## wherein:
Y is cyano or --C(O)R.sub.a, further wherein: R.sub.a is: hydrogen,
hydroxy, halo, amino, azido, mercapto or silyl; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
alkoxyamino.sub.(C.ltoreq.12), dialkylamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12), or a
substituted version of any of these groups;
X is OR.sub.b, NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and
R.sub.c are each independently: hydrogen or hydroxy;
alkyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or a substituent
convertible in vivo to hydrogen; provided that R.sub.b is absent
when the atom to which it is bound is part of a double bond,
further provided that when R.sub.b is absent the atom to which it
is bound is part of a double bond;
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and
R.sub.4 is alkyl.sub.(C.ltoreq.8) or substituted
alkyl.sub.(C.ltoreq.8);
or pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00006## wherein:
R.sub.a is: hydrogen, hydroxy, halo, or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
alkenyloxy.sub.(C.ltoreq.8), alkynyloxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), aralkoxy.sub.(C.ltoreq.8),
heteroaryloxy.sub.(C.ltoreq.8), heteroaralkoxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
alkoxyamino.sub.(C.ltoreq.8), dialkylamino.sub.(C.ltoreq.8),
alkenylamino.sub.(C.ltoreq.8), alkynylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), aralkylamino.sub.(C.ltoreq.8),
heteroarylamino.sub.(C.ltoreq.8),
heteroaralkylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
X is OR.sub.b or NR.sub.bR.sub.c, wherein R.sub.b and R.sub.c are
each independently: hydrogen or hydroxy; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), acyloxy.sub.(C.ltoreq.8),
alkylamino.sub.(C.ltoreq.8), arylamino.sub.(C.ltoreq.8),
amido.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.b is absent when the atom to which it is bound is part
of a double bond, further provided that when R.sub.b is absent the
atom to which it is bound is part of a double bond; and
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or pharmaceutically
acceptable salts, esters, hydrates, solvates, tautomers, prodrugs,
or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00007## wherein:
R.sub.a is: hydrogen, hydroxy, halo, or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
alkenyloxy.sub.(C.ltoreq.8), alkynyloxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), aralkoxy.sub.(C.ltoreq.8),
heteroaryloxy.sub.(C.ltoreq.8), heteroaralkoxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
alkoxyamino.sub.(C.ltoreq.8), dialkylamino.sub.(C.ltoreq.8),
alkenylamino.sub.(C.ltoreq.8), alkynylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), aralkylamino.sub.(C.ltoreq.8),
heteroarylamino.sub.(C.ltoreq.8),
heteroaralkylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and
R.sub.2 is: cyano or fluoro; or fluoroalkyl.sub.(C.ltoreq.5),
alkenyl.sub.(C.ltoreq.5), alkynyl.sub.(C.ltoreq.5),
heteroaryl.sub.(C.ltoreq.5), acyl.sub.(C.ltoreq.5),
acyloxy.sub.(C.ltoreq.5), amido.sub.(C.ltoreq.5), or a substituted
version of any of these groups; or pharmaceutically acceptable
salts, esters, hydrates, solvates, tautomers, prodrugs, or optical
isomers thereof.
In some embodiments, the compound is further defined as:
##STR00008## wherein R.sub.a is: hydrogen, hydroxy, halo or amino;
or alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), alkoxy.sub.(C.ltoreq.8),
alkenyloxy.sub.(C.ltoreq.8), alkynyloxy.sub.(C.ltoreq.8),
aryloxy.sub.(C.ltoreq.8), aralkoxy.sub.(C.ltoreq.8),
heteroaryloxy.sub.(C.ltoreq.8), heteroaralkoxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
alkoxyamino.sub.(C.ltoreq.8), dialkylamino.sub.(C.ltoreq.8),
alkenylamino.sub.(C.ltoreq.8), alkynylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), aralkylamino.sub.(C.ltoreq.8),
heteroarylamino.sub.(C.ltoreq.8),
heteroaralkylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or pharmaceutically
acceptable salts, esters, hydrates, solvates, tautomers, prodrugs,
or optical isomers thereof
In some embodiments, the compound is further defined as:
##STR00009## wherein R.sub.a is alkoxy.sub.(C1-4),
alkylamino.sub.(C1-4), alkoxyamino.sub.(C1-4),
dialkylamino.sub.(C2-4), or a substituted version of any of these
groups; or pharmaceutically acceptable salts, esters, hydrates,
solvates, tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00010## wherein R.sub.a is alkyl.sub.(C1-4) or
aralkoxy.sub.(C7-8) or a substituted version of either of these
groups; or pharmaceutically acceptable salts, esters, hydrates,
solvates, tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00011## wherein R.sub.a is hydrogen, hydroxy, amino,
dimethylamino, methyl, methoxy, methoxyamino, benzyloxy, or
2,2,2-trifluoroethylamino; or pharmaceutically acceptable salts,
hydrates, solvates, tautomers, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00012## wherein R.sub.a is hydrogen, hydroxy, amino, methoxy,
or 2,2,2-trifluoroethylamino; or pharmaceutically acceptable salts,
hydrates, solvates, tautomers, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00013## wherein R.sub.d is alkyl.sub.(C.ltoreq.8),
alkenyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), or a substituted version of any of
these groups; or pharmaceutically acceptable salts, hydrates,
solvates, tautomers, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00014## wherein Y is heteroaryl.sub.(C.ltoreq.8) or a
substituted heteroaryl.sub.(C.ltoreq.8); or a pharmaceutically
acceptable salts, hydrates, solvates, tautomers, or optical isomers
thereof.
In some embodiments, the compound is further defined as:
##STR00015## wherein:
wherein Y is cyano or --C(O)R.sub.a, further wherein: R.sub.a is:
hydrogen, hydroxy, halo, amino, azido, mercapto or silyl; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
alkoxyamino.sub.(C.ltoreq.12), dialkylamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.3 is: absent or hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.3 is absent when the oxygen atom to which it is bound is
part of a double bond, further provided that when R.sub.3 is absent
the oxygen atom to which it is bound is part of a double bond;
and
R.sub.4 is alkyl.sub.(C.ltoreq.8) or substituted
alkyl.sub.(C.ltoreq.8);
or pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00016## wherein:
Y is cyano or --C(O)R.sub.a, wherein R.sub.a is: hydrogen, hydroxy,
halo, amino, hydroxyamino, azido or mercapto; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
alkoxyamino.sub.(C.ltoreq.12), dialkylamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12),
alkylsulfonylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and
R.sub.3 is: hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; or
pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00017## wherein:
X is OR.sub.b, NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and
R.sub.c are each independently: hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.b is absent when the atom to which it is bound is part
of a double bond, further provided that when R.sub.b is absent the
atom to which it is bound is part of a double bond;
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and
R.sub.4 is alkyl.sub.(C.ltoreq.8) or substituted
alkyl.sub.(C.ltoreq.8);
or pharmaceutically acceptable salts, esters, hydrates, solvates,
tautomers, prodrugs, or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00018## wherein:
X is OR.sub.b or NR.sub.bR.sub.c, wherein R.sub.b and R.sub.c are
each independently: hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or a substituent convertible in vivo to hydrogen; provided
that R.sub.b is absent when the atom to which it is bound is part
of a double bond, further provided that when R.sub.b is absent the
atom to which it is bound is part of a double bond; and
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or pharmaceutically
acceptable salts, esters, hydrates, solvates, tautomers, prodrugs,
or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00019## wherein:
R.sub.a is: hydrogen, hydroxy, halo, amino, azido, mercapto or
silyl; or alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
alkoxyamino.sub.(C.ltoreq.12), dialkylamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12), or a
substituted version of any of these groups;
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups;
R.sub.2 is: cyano, hydroxy, halo or amino; or
fluoroalkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
heteroaryl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; and or pharmaceutically
acceptable salts, esters, hydrates, solvates, tautomers, prodrugs,
or optical isomers thereof.
In some embodiments, the compound is further defined as:
##STR00020## wherein R.sub.a is: hydrogen, hydroxy, halo or amino;
or alkyl.sub.(C.ltoreq.6), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.6), aryloxy.sub.(C.ltoreq.8),
aralkoxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.6),
alkoxyamino.sub.(C.ltoreq.6), alkoxyamino.sub.(C.ltoreq.6),
dialkylamino.sub.(C.ltoreq.6), arylamino.sub.(C.ltoreq.8),
aralkylamino.sub.(C.ltoreq.8), heteroarylamino.sub.(C.ltoreq.6),
heteroarylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.6), or a
substituted version of any of these groups; or pharmaceutically
acceptable salts, esters, hydrates, solvates, tautomers, prodrugs,
or optical isomers thereof.
In some aspects, the disclosure provides compounds of the
formula:
##STR00021## wherein:
Y is cyano or --C(O)R.sub.a, wherein R.sub.a is: hydrogen, hydroxy,
halo, amino, hydroxyamino, azido or mercapto; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkynyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), aralkoxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
dialkylamino.sub.(C.ltoreq.12), alkoxyamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12),
alkylsulfonylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12),
alkylthio.sub.(C.ltoreq.12), alkenylthio.sub.(C.ltoreq.12),
alkynylthio.sub.(C.ltoreq.12), arylthio.sub.(C.ltoreq.12),
aralkylthio.sub.(C.ltoreq.12), heteroarylthio.sub.(C.ltoreq.12),
heteroaralkylthio.sub.(C.ltoreq.12), acylthio.sub.(C.ltoreq.12),
alkylammonium.sub.(C.ltoreq.12), alkylsulfonium.sub.(C.ltoreq.12),
alkylsilyl.sub.(C.ltoreq.12), or a substituted version of any of
these groups;
X is OR.sub.b, NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and
R.sub.c are each independently: hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or provided that R.sub.b is absent when the atom to which
it is bound is part of a double bond, further provided that when
R.sub.b is absent the atom to which it is bound is part of a double
bond; and
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; or salts, esters,
hydrates, solvates, tautomers, or optical isomers thereof.
In some aspects, the disclosure provides compounds of the
formula:
##STR00022## wherein:
Y is cyano or --C(O)R.sub.a, wherein R.sub.a is: hydrogen, hydroxy,
halo, amino, hydroxyamino, azido or mercapto; or
alkyl.sub.(C.ltoreq.12), alkenyl.sub.(C.ltoreq.12),
alkylnyl.sub.(C.ltoreq.12), aryl.sub.(C.ltoreq.12),
aralkyl.sub.(C.ltoreq.12), heteroaryl.sub.(C.ltoreq.12),
heteroaralkyl.sub.(C.ltoreq.12), alkoxy.sub.(C.ltoreq.12),
alkenyloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), alkynyloxy.sub.(C.ltoreq.12),
heteroaryloxy.sub.(C.ltoreq.12), heteroaralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), alkylamino.sub.(C.ltoreq.12),
dialkylamino.sub.(C.ltoreq.12), alkoxyamino.sub.(C.ltoreq.12),
alkenylamino.sub.(C.ltoreq.12), alkynylamino.sub.(C.ltoreq.12),
arylamino.sub.(C.ltoreq.12), aralkylamino.sub.(C.ltoreq.12),
heteroarylamino.sub.(C.ltoreq.12),
heteroaralkylamino.sub.(C.ltoreq.12),
alkylsulfonylamino.sub.(C.ltoreq.12), amido.sub.(C.ltoreq.12),
alkylthio.sub.(C.ltoreq.12), alkenylthio.sub.(C.ltoreq.12),
alkynylthio.sub.(C.ltoreq.12), arylthio.sub.(C.ltoreq.12),
aralkylthio.sub.(C.ltoreq.12), heteroarylthio.sub.(C.ltoreq.12),
heteroaralkylthio.sub.(C.ltoreq.12), acylthio.sub.(C.ltoreq.12),
alkylammonium.sub.(C.ltoreq.12), alkylsulfonium.sub.(C.ltoreq.12),
alkylsilyl.sub.(C.ltoreq.12), or a substituted version of any of
these groups;
X is OR.sub.b, NR.sub.bR.sub.c, or SR.sub.b, wherein R.sub.b and
R.sub.c are each independently: hydrogen; alkyl.sub.(C.ltoreq.8),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
acyl.sub.(C.ltoreq.8), or a substituted version of any of these
groups; or provided that R.sub.b is absent when the atom to which
it is bound is part of a double bond, further provided that when
R.sub.b is absent the atom to which it is bound is part of a double
bond; and
R.sub.1 is: hydrogen, cyano, hydroxy, halo or amino; or
alkyl.sub.(C.ltoreq.8), alkenyl.sub.(C.ltoreq.8),
alkynyl.sub.(C.ltoreq.8), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaryl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), acyl.sub.(C.ltoreq.8),
alkoxy.sub.(C.ltoreq.8), aryloxy.sub.(C.ltoreq.8),
acyloxy.sub.(C.ltoreq.8), alkylamino.sub.(C.ltoreq.8),
arylamino.sub.(C.ltoreq.8), amido.sub.(C.ltoreq.8), or a
substituted version of any of these groups; R' is hydroxy,
alkoxy.sub.(C.ltoreq.12), substituted alkoxy.sub.(C.ltoreq.12),
aryloxy.sub.(C.ltoreq.12), substituted aryloxy.sub.(C.ltoreq.12),
aralkoxy.sub.(C.ltoreq.12), substituted aralkoxy.sub.(C.ltoreq.12),
acyloxy.sub.(C.ltoreq.12), or substituted
acyloxy.sub.(C.ltoreq.12); or salts, esters, hydrates, solvates,
tautomers, or optical isomers thereof.
In a variation of each of the above embodiments containing a Z
group, Z can be a single bond, --O--, or --NH--. In a variation of
each of the above embodiments containing an X group, X can be
OR.sub.b. In some variations, R.sub.b is absent. In other
variations, R.sub.b is hydrogen. In other variations, X can be
NR.sub.b. In some variations, R.sub.b can be hydroxy. In a
variation of each of the above embodiments containing a Y group, Y
can be cyano or --C(O)R.sub.a. In some variations, R.sub.a can be
hydroxy. In some variations, R.sub.a can be
alkoxy.sub.(C.ltoreq.6), aryloxy.sub.(C.ltoreq.8),
aralkyloxy.sub.(C.ltoreq.8), or a substituted version of any of
these groups. In some of these variations, R.sub.a can be
alkoxy.sub.(C2-6). In some of these variations, R.sub.a can be
alkoxy.sub.(C1-5) or substituted alkoxy.sub.(C1-5). In some of
these variations, R.sub.a can be alkoxy.sub.(C2-4) or substituted
alkoxy.sub.(C2-4). In some of these variations, R.sub.a can be
alkoxy.sub.(C1-4) or substituted alkoxy.sub.(C1-4). In some of
these variations, R.sub.a can be alkoxy.sub.(C1-2) or substituted
alkoxy.sub.(C1-2). For example, R.sub.a can be methoxy. In some
variations, R.sub.a can be amino. In some variations, R.sub.a can
be alkylamino.sub.(C1-6), alkoxyamino.sub.(C1-6),
arylamino.sub.(C1-8), aralkylamino.sub.(C1-8),
dialkylamino.sub.(C2-8), or a substituted version of any of these
groups. In some of these variations, R.sub.a can be
alkylamino.sub.(C2-6) or substituted alkylamino.sub.(C2-6). In some
of these variations, R.sub.a can be alkylamino.sub.(C3-6). In some
variations, R.sub.a can be alkylamino.sub.(C1-5),
dialkylamino.sub.(C2-6), or substituted version of either of these
groups. In some variations, R.sub.a can be alkylamino.sub.(C2-4),
dialkylamino.sub.(C2-5), or substituted version of either of these
groups. In some variations, R.sub.a can be alkylamino.sub.(C1-4) or
substituted alkylamino.sub.(C1-4). In some variations, R.sub.a can
be alkylamino.sub.(C1-3). In some of these variations, R.sub.a can
be methylamino or ethylamino. In some of these variations, R.sub.a
can be substituted alkylamino.sub.(C1-3). For example, R.sub.a can
be 2,2,2-trifluoroethylamino. In some of these variations, R.sub.a
can be alkyl.sub.(C1-5), aryl.sub.(C.ltoreq.8),
aralkyl.sub.(C.ltoreq.8), heteroaralkyl.sub.(C.ltoreq.8), or a
substituted version of any of these groups. In some of these
variations, R.sub.a can be heteroaryl.sub.(C1-8) or substituted
heteroaryl.sub.(C1-8). For example, wherein R.sub.a can be
imidazolyl. In some of these variations, R.sub.a can be --H.
In a variation of each of the above embodiments containing an
R.sub.1 group, R.sub.1 can be --H, --OH or --F. For example,
R.sub.1 can be --H. In a variation of each of the above embodiments
containing an R.sub.2 group, R.sub.2 can be --CN. In some
variations, R.sub.2 can be a substituted acyl.sub.(C1-3), such as
--C(.dbd.O)NHS(.dbd.O).sub.2CH.sub.3. In some variations, R.sub.2
is fluoroalkyl.sub.(C.ltoreq.8). For example, R.sub.2 can be
--CF.sub.3. In other variations, R.sub.2 is not
fluoroalkyl.sub.(C.ltoreq.8).
In a variation of each of the above embodiments containing an
R.sub.3 group, R.sub.3 can be hydrogen or acetyl. In another
variation, R.sub.3 can be absent. In a variation of each of the
above embodiments containing an R.sub.4 group, R.sub.4 can be
methyl or hydroxymethyl. In a variation of each of the above
embodiments containing an R.sub.6, R.sub.7, R.sub.8, or R.sub.9
group, R.sub.6, R.sub.7, R.sub.8, or R.sub.9 can independently be
hydrogen. In a variation of each of the above embodiments
containing an R.sub.10 or R.sub.11 group, R.sub.10 or R.sub.11 can
independently be methyl. In a variation of each of the above
embodiments containing an R' group, R' can be acetyloxy or
hydroxy.
In a variation of each of the above embodiments containing an
R.sub.d group, R.sub.d can be alkyl.sub.(C1-5),
aryl.sub.(C.ltoreq.8), aralkyl.sub.(C.ltoreq.8),
heteroaralkyl.sub.(C.ltoreq.8), or a substituted version of any of
these groups. In some variations, R.sub.d can be alkyl.sub.(C1-4)
or a substituted version thereof. In some variations, R.sub.d can
be alkyl.sub.(C1-3) or a substituted version thereof. In some
variations, R.sub.d can be alkyl.sub.(C1-2) or a substituted
version thereof.
Non-limiting examples of compounds provided by this invention
include the compounds according to the formulas shown below, as
well as or pharmaceutically acceptable salts thereof. In certain
embodiments, these compounds are substantially free from other
optical isomers thereof
##STR00023## ##STR00024## ##STR00025## ##STR00026##
##STR00027##
Examples of specific compounds provided by the present disclosure
include: (4aS,6aR,6bR,8aR,12aR,14aR,14bS)-methyl
11-cyano-2,2,6a,6b,9,9,12a-heptamethyl-10,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7-
,8,8a,9,10,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxylate,
(4aS,6aR,6bR,8aR,12aR,14aR,14bS)-methyl
11-cyano-10-hydroxy-2,2,6a,6b,9,9,12a-heptamethyl-14-oxo-1,2,3,4,4a,5,6,6-
a,6b,7,8,8a,9,12,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxylate,
(4aS,6aR,6bR,8aR,12aR,14aR,14bS)-11-cyano-2,2,6a,6b,9,9,12a-heptamethyl-1-
0,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b-icosahyd-
ropicene-4a-carboxylic acid,
(4aR,6aR,6bR,8aS,12aS,12bR,14bR)-4,4,6a,6b,11,11,14b-heptamethyl-3,13-dio-
xo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11,12,12a,12b,13,14,14a,14b-icosahydropice-
ne-2,8a-dicarbonitrile,
(4aS,6aR,6bR,8aR,12aR,14aR,14bS)-11-cyano-2,2,6a,6b,9,9,12a-heptamethyl-1-
0,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b-icosahyd-
ropicene-4a-carboxamide,
(4aR,6aR,6bR,8aS,12aS,12bR,14bR)-8a-formyl-4,4,6a,6b,11,11,14b-heptamethy-
l-3,13-dioxo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11,12,12a,12b,13,14,14a,14b-icos-
ahydropicene-2-carbonitrile,
(4aS,6aR,6bR,8aR,12aR,14R,14aR,14bS)-methyl
11-cyano-14-hydroxy-2,2,6a,6b,9,9,12a-heptamethyl-10-oxo-1,2,3,4,4a,5,6,6-
a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxylate,
(4aS,6aR,6bR,8aR,12aR,14aR,14bS)-11-cyano-2,2,6a,6b,9,9,12a-heptamethyl-1-
0,14-dioxo-N-(2,2,2-trifluoroethyl)-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,1-
2b,13,14,14a,14b-icosahydropicene-4a-carboxamide,
(4aS,6aR,6bR,8aR,12aR,12bR,14R,14aR,14bS)-2,2,2-trifluoroethyl
11-cyano-14-hydroxy-2,2,6a,6b,9,9,12a-heptamethyl-10-oxo-1,2,3,4,4a,5,6,6-
a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxylate,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-11-cyano-2,2,6a,6b,9,9,12a-heptamet-
hyl-10,14-dioxo-N-(2,2,2-trifluoroethyl)-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,-
12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxamide,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)--N'-acetyl-11-cyano-2,2,6a,6b,9,9,1-
2a-heptamethyl-10,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,-
14a,14b-icosahydropicene-4a-carbohydrazide,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-11-cyano-2,2,6a,6b,9,9,12a-heptamet-
hyl-10,14-dioxo-N-(2,2,2-trifluoroethyl)docosahydropicene-4a-carboxamide,
(4aR,6aR,6bR,8aS,12aS,12bR,14aR,14bR)-4,4,6a,6b,11,11,14b-heptamethyl-3,1-
3-dioxo-8a-(2H-tetrazol-5-yl)-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11,12,12a,12b,1-
3,14,14a,14b-icosahydropicene-2-carbonitrile,
(4aR,6aR,6bR,8aS,12aS,12bR,14aR,14bR)-4,4,6a,6b,11,11,14b-heptamethyl-8a--
(2-methyl-2H-tetrazol-5-yl)-3,13-dioxo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11,12,-
12a,12b,13,14,14a,14b-icosahydropicene-2-carbonitrile,
(4aR,6aR,6bR,8aS,12aS,12bR,14aR,14bR)-4,4,6a,6b,11,11,14b-heptamethyl-8a--
(5-methyl-1,3,4-oxadiazol-2-yl)-3,13-dioxo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11-
,12,12a,12b,13,14,14a,14b-icosahydropicene-2-carbonitrile, and
(4aR,6aR,6bR,8aS,12aS,12bR,14aR,14bR)-4,4,6a,6b,11,11,14b-heptamethyl-8a--
(5-methyl-1,3,4-thiadiazol-2-yl)-3,13-dioxo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,1-
1,12,12a,12b,13,14,14a,14b-icosahydropicene-2-carbonitrile,
(4aR,6aR,6bR,8aS,12aS,12bR,14aR,14bR)-8a-acetyl-4,4,6a,6b,11,11,14b-hepta-
methyl-3,13-dioxo-3,4,4a,5,6,6a,6b,7,8,8a,9,10,11,12,12a,12b,13,14,14a,14b-
-icosahydropicene-2-carbonitrile,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-benzyl
11-cyano-2,2,6a,6b,9,9,12a-heptamethyl-10,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7-
,8,8a,9,10,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxylate,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-11-cyano-N,N,2,2,6a,6b,9,9,12a-nona-
methyl-10,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b--
icosahydropicene-4a-carboxamide,
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-11-cyano-N-methoxy-2,2,6a,6b,9,9,12-
a-heptamethyl-10,14-dioxo-1,2,3,4,4a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,1-
4a,14b-icosahydropicene-4a-carboxamide
(4aS,6aR,6bR,8aR,12aR,12bR,14aR,14bS)-methyl
11-cyano-14-(hydroxyimino)-2,2,6a,6b,9,9,12a-heptamethyl-10-oxo-1,2,3,4,4-
a,5,6,6a,6b,7,8,8a,9,10,12a,12b,13,14,14a,14b-icosahydropicene-4a-carboxyl-
ate, (4 aR,6aR,6bS,8aS,12aR,15aR,15bR)-methyl
2-cyano-14-hydroxy-4,4,6a,6b,11,11,15b-heptamethyl-3-oxo-3,4,4a,5,6,6a,6b-
,7,8,8a,9,10,11,12,12a,12b,14,15,15a,15b-icosahydrodinaphtho[1,2-b:2',1'-d-
]oxepine-8a-carboxylate, and
(4aR,6aR,6bS,8aS,12aR,12bR,15aR,15bR)-methyl
2-cyano-4,4,6a,6b,11,11,15b-heptamethyl-3,14-dioxo-4,4a,5,6,6a,6b,7,8,8a,-
9,10,11,12,12a,12b,13,14,15,15a,15b-icosahydro-3H-dinaphtho[1,2-b:2',1'-d]-
azepine-8a-carboxylate.
In some embodiments, compounds of the present disclosure are in the
form of pharmaceutically acceptable salts. In other embodiments,
compounds of the present disclosure are not in the form of a
pharmaceutically acceptable salts. In some embodiments, compounds
of the present disclosure are in the form of a hydrate. In other
embodiments, compounds of the present disclosure are not in the
form of a hydrate. In some embodiments, compounds of the present
disclosure are in the form of a solvate. In other embodiments,
compounds of the present disclosure are not in the form of a
solvate.
In some embodiments, compounds of the present disclosure can be
esters of the above formulas. The ester may, for example, result
from a condensation reaction between a hydroxy group of the formula
and the carboxylic acid group of biotin. In other embodiments,
compounds of the present disclosure are not an ester.
In some embodiments, the compounds of the present disclosure can be
present as a mixture of stereoisomers. In other embodiments, the
compounds of the present disclosure are present as single
stereoisomers.
In some embodiments, compounds of the present disclosure may be
inhibitors of IFN-.gamma.-induced nitrous oxide (NO) production in
macrophages, for example, having an IC.sub.50 value of less than
0.2 .mu.M.
Other general aspects of the present disclosure contemplate a
pharmaceutical composition comprising as an active ingredient a
compound of the present disclosure and a pharmaceutically
acceptable carrier. The composition may, for example, be adapted
for administration by a route selected from the group consisting of
orally, intraadiposally, intraarterially, intraarticularly,
intracranially, intradermally, intralesionally, intramuscularly,
intranasally, intraocularally, intrapericardially,
intraperitoneally, intrapleurally, intraprostaticaly,
intrarectally, intrathecally, intratracheally, intratumorally,
intraumbilically, intravaginally, intravenously, intravesicularlly,
intravitreally, liposomally, locally, mucosally, orally,
parenterally, rectally, subconjunctival, subcutaneously,
sublingually, topically, transbuccally, transdermally, vaginally,
in cremes, in lipid compositions, via a catheter, via a lavage, via
continuous infusion, via infusion, via inhalation, via injection,
via local delivery, via localized perfusion, bathing target cells
directly, or any combination thereof. In particular embodiments,
the composition may be formulated for oral delivery. In particular
embodiments, the composition is formulated as a hard or soft
capsule, a tablet, a syrup, a suspension, a wafer, or an elixir. In
certain embodiments, the soft capsule is a gelatin capsule. Certain
compositions may comprise a protective coating, such as those
compositions formulated for oral delivery. Certain compositions
further comprise an agent that delays absorption, such as those
compositions formulated for oral delivery. Certain compositions may
further comprise an agent that enhances solubility or
dispersibility, such as those compositions formulated for oral
delivery. Certain compositions may comprise a compound of the
present disclosure, wherein the compound is dispersed in a
liposome, an oil in water emulsion or a water in oil emulsion.
Yet another general aspect of the present disclosure contemplates a
therapeutic method comprising administering a pharmaceutically
effective compound of the present disclosure to a subject. The
subject may, for example, be a human. These or any other methods of
the present disclosure may further comprise identifying a subject
in need of treatment.
Another method of the present disclosure contemplates a method of
treating cancer in a subject, comprising administering to the
subject a pharmaceutically effective amount of a compound of the
present disclosure. The cancer may be any type of cancer, such as a
carcinoma, sarcoma, lymphoma, leukemia, melanoma, mesothelioma,
multiple myeloma, or seminoma. Other types of cancers include
cancer of the bladder, blood, bone, brain, breast, central nervous
system, colon, endometrium, esophagus, genitourinary tract, head,
larynx, liver, lung, neck, ovary, pancreas, prostate, spleen, small
intestine, large intestine, stomach, or testicle. In these or any
other methods, the subject may be a primate. In these or any other
methods, the subject may be a human. This or any other method may
further comprise identifying a subject in need of treatment. The
subject may have a family or patient history of cancer. In certain
embodiments, the subject has symptoms of cancer. The compounds of
the invention may be administered via any method described herein,
such as locally. In certain embodiments, the compound is
administered by direct intratumoral injection or by injection into
tumor vasculature. In certain embodiments, the compounds may be
administered systemically. The compounds may be administered
intravenously, intra-arterially, intramuscularly,
intraperitoneally, subcutaneously or orally, in certain
embodiments.
In certain embodiments regarding methods of treating cancer in a
subject, comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure, the
pharmaceutically effective amount is 0.1-1000 mg/kg. In certain
embodiments, the pharmaceutically effective amount is administered
in a single dose per day. In certain embodiments, the
pharmaceutically effective amount is administered in two or more
doses per day. The compound may be administered by contacting a
tumor cell during ex vivo purging, for example. The method of
treatment may comprise any one or more of the following: a)
inducing cytotoxicity in a tumor cell; b) killing a tumor cell; c)
inducing apoptosis in a tumor cell; d) inducing differentiation in
a tumor cell; or e) inhibiting growth in a tumor cell. The tumor
cell may be any type of tumor cell, such as a leukemia cell. Other
types of cells include, for example, a bladder cancer cell, a
breast cancer cell, a lung cancer cell, a colon cancer cell, a
prostate cancer cell, a liver cancer cell, a pancreatic cancer
cell, a stomach cancer cell, a testicular cancer cell, a brain
cancer cell, an ovarian cancer cell, a lymphatic cancer cell, a
skin cancer cell, a brain cancer cell, a bone cancer cell, or a
soft tissue cancer cell.
Combination treatment therapy is also contemplated by the present
disclosure. For example, regarding methods of treating cancer in a
subject, comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure, the
method may further comprise a treatment selected from the group
consisting of administering a pharmaceutically effective amount of
a second drug, radiotherapy, gene therapy, and surgery. Such
methods may further comprise (1) contacting a tumor cell with the
compound prior to contacting the tumor cell with the second drug,
(2) contacting a tumor cell with the second drug prior to
contacting the tumor cell with the compound, or (3) contacting a
tumor cell with the compound and the second drug at the same time.
The second drug may, in certain embodiments, be an antibiotic,
anti-inflammatory, anti-neoplastic, anti-proliferative, anti-viral,
immunomodulatory, or immunosuppressive. The second drug may be an
alkylating agent, androgen receptor modulator, cytoskeletal
disruptor, estrogen receptor modulator, histone-deacetylase
inhibitor, HMG-CoA reductase inhibitor, prenyl-protein transferase
inhibitor, retinoid receptor modulator, topoisomerase inhibitor, or
tyrosine kinase inhibitor. In certain embodiments, the second drug
is 5-azacitidine, 5-fluorouracil, 9-cis-retinoic acid, actinomycin
D, alitretinoin, all-trans-retinoic acid, annamycin, axitinib,
belinostat, bevacizumab, bexarotene, bosutinib, busulfan,
capecitabine, carboplatin, carmustine, CD437, cediranib, cetuximab,
chlorambucil, cisplatin, cyclophosphamide, cytarabine, dacarbazine,
dasatinib, daunorubicin, decitabine, docetaxel, dolastatin-10,
doxifluridine, doxorubicin, doxorubicin, epirubicin, erlotinib,
etoposide, etoposide, gefitinib, gemcitabine, gemtuzumab
ozogamicin, hexamethylmelamine, idarubicin, ifosfamide, imatinib,
irinotecan, isotretinoin, ixabepilone, lapatinib, LBH589,
lomustine, mechlorethamine, melphalan, mercaptopurine,
methotrexate, mitomycin, mitoxantrone, MS-275, neratinib,
nilotinib, nitrosourea, oxaliplatin, paclitaxel, plicamycin,
procarbazine, semaxanib, semustine, sodium butyrate, sodium
phenylacetate, streptozotocin, suberoylanilide hydroxamic acid,
sunitinib, tamoxifen, teniposide, thiopeta, tioguanine, topotecan,
TRAIL, trastuzumab, tretinoin, trichostatin A, valproic acid,
valrubicin, vandetanib, vinblastine, vincristine, vindesine, or
vinorelbine.
Methods of treating or preventing a disease with an inflammatory
component in a subject, comprising administering to the subject a
pharmaceutically effective amount of a compound of the present
disclosure are also contemplated. The disease may be, for example,
lupus or rheumatoid arthritis. The disease may be an inflammatory
bowel disease, such as Crohn's disease or ulcerative colitis. The
disease with an inflammatory component may be a cardiovascular
disease. The disease with an inflammatory component may be
diabetes, such as type 1 or type 2 diabetes. Compounds of the
present disclosure may also be used to treat complications
associated with diabetes. Such complications are well-known in the
art and include, for example, obesity, hypertension,
atherosclerosis, coronary heart disease, stroke, peripheral
vascular disease, hypertension, nephropathy, neuropathy,
myonecrosis, retinopathy and metabolic syndrome (syndrome X). The
disease with an inflammatory component may be a skin disease, such
as psoriasis, acne, or atopic dermatitis. Administration of a
compound of the present disclosure in treatment methods of such
skin diseases may be, for example, topical or oral.
The disease with an inflammatory component may be metabolic
syndrome (syndrome X). A patient having this syndrome is
characterized as having three or more symptoms selected from the
following group of five symptoms: (1) abdominal obesity; (2)
hypertriglyceridemia; (3) low high-density lipoprotein cholesterol
(HDL); (4) high blood pressure; and (5) elevated fasting glucose,
which may be in the range characteristic of Type 2 diabetes if the
patient is also diabetic. Each of these symptoms is defined in the
Third Report of the National Cholesterol Education Program Expert
Panel on Detection, Evaluation and Treatment of High Blood
Cholesterol in Adults (Adult Treatment Panel III, or ATP III),
National Institutes of Health, 2001, NIH Publication No. 01-3670,
incorporated herein by reference. Patients with metabolic syndrome,
whether or not they have or develop overt diabetes mellitus, have
an increased risk of developing the macrovascular and microvascular
complications that are listed above that occur with type 2
diabetes, such as atherosclerosis and coronary heart disease.
Another general method of the present disclosure entails a method
of treating or preventing a cardiovascular disease in a subject,
comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure. The
cardiovascular disease may be, for example, atherosclerosis,
cardiomyopathy, congenital heart disease, congestive heart failure,
myocarditis, rheumatic heart disease, valve disease, coronary
artery disease, endocarditis, or myocardial infarction. Combination
therapy is also contemplated for such methods. For example, such
methods may further comprise administering a pharmaceutically
effective amount of a second drug. The second drug may be, for
example, a cholesterol lowering drug, an anti-hyperlipidemic, a
calcium channel blocker, an anti-hypertensive, or an HMG-CoA
reductase inhibitor. Non-limiting examples of second drugs include
amlodipine, aspirin, ezetimibe, felodipine, lacidipine,
lercanidipine, nicardipine, nifedipine, nimodipine, nisoldipine or
nitrendipine. Other non-limiting examples of second drugs include
atenolol, bucindolol, carvedilol, clonidine, doxazosin, indoramin,
labetalol, methyldopa, metoprolol, nadolol, oxprenolol,
phenoxybenzamine, phentolamine, pindolol, prazosin, propranolol,
terazosin, timolol or tolazoline. The second drug may be, for
example, a statin, such as atorvastatin, cerivastatin, fluvastatin,
lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin or
simvastatin.
Methods of treating or preventing a neurodegenerative disease in a
subject, comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure are also
contemplated. The neurodegenerative disease may, for example, be
selected from the group consisting of Parkinson's disease,
Alzheimer's disease, multiple sclerosis (MS), Huntington's disease
and amyotrophic lateral sclerosis. In particular embodiments, the
neurodegenerative disease is Alzheimer's disease. In particular
embodiments, the neurodegenerative disease is MS, such as primary
progressive, relapsing-remitting secondary progressive or
progressive relapsing MS. The subject may be, for example, a
primate. The subject may be a human.
In particular embodiments of methods of treating or preventing a
neurodegenerative disease in a subject, comprising administering to
the subject a pharmaceutically effective amount of a compound of
the present disclosure, the treatment suppresses the demyelination
of neurons in the subject's brain or spinal cord. In certain
embodiments, the treatment suppresses inflammatory demyelination.
In certain embodiments, the treatment suppresses the transection of
neuron axons in the subject's brain or spinal cord. In certain
embodiments, the treatment suppresses the transection of neurites
in the subject's brain or spinal cord. In certain embodiments, the
treatment suppresses neuronal apoptosis in the subject's brain or
spinal cord. In certain embodiments, the treatment stimulates the
remyelination of neuron axons in the subject's brain or spinal
cord. In certain embodiments, the treatment restores lost function
after an MS attack. In certain embodiments, the treatment prevents
a new MS attack. In certain embodiments, the treatment prevents a
disability resulting from an MS attack.
One general aspect of the present disclosure contemplates a method
of treating or preventing a disorder characterized by
overexpression of iNOS genes in a subject, comprising administering
to the subject a pharmaceutically effective amount of a compound of
the present disclosure.
Another general aspect of the present disclosure contemplates a
method of inhibiting IFN-.gamma.-induced nitric oxide production in
cells of a subject, comprising administering to said subject a
pharmaceutically effective amount of a compound of the present
disclosure.
Yet another general method of the present disclosure contemplates a
method of treating or preventing a disorder characterized by
overexpression of COX-2 genes in a subject, comprising
administering to the subject a pharmaceutically effective amount of
compound of the present disclosure.
Methods of treating renal/kidney disease (RKD) in a subject,
comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure are also
contemplated. See U.S. patent application Ser. No. 12/352,473,
which is incorporated by reference herein in its entirety. The RKD
may result from, for example, a toxic insult. The toxic insult may
result from, for example, an imaging agent or a drug. The drug may
be a chemotherapeutic, for example. The RKD may result from
ischemia/reperfusion injury, in certain embodiments. In certain
embodiments, the RKD results from diabetes or hypertension. The RKD
may result from an autoimmune disease. The RKD may be further
defined as chronic RKD, or acute RKD.
In certain methods of treating renal/kidney disease (RKD) in a
subject, comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure, the
subject has undergone or is undergoing dialysis. In certain
embodiments, the subject has undergone or is a candidate to undergo
kidney transplant. The subject may be a primate. The primate may be
a human. The subject in this or any other method may be, for
example, a cow, horse, dog, cat, pig, mouse, rat or guinea pig.
Also contemplated by the present disclosure is a method for
improving glomerular filtration rate or creatinine clearance in a
subject, comprising administering to the subject a pharmaceutically
effective amount of a compound of the present disclosure.
In some embodiments, the invention provides compounds useful for
preventing and/or treating diseases or disorders whose pathology
involves oxidative stress, inflammation, and/or dysregulation of
inflammatory signaling pathways. In some variations, the diseases
or disorders can be characterized by overexpression of inducible
nitric oxide synthase (iNOS) and/or inducible cyclooxygenase
(COX-2) in affected tissues. In some variations, the diseases or
disorders can be characterized by overproduction of reactive oxygen
species (ROS) or reactive nitrogen species (RNS) such as
superoxide, hydrogen peroxide, nitric oxide or peroxynitrite in
affected tissues. In some variations, the disease or disorder is
characterized by excessive production of inflammatory cytokines or
other inflammation-related proteins such as TNF.alpha., IL-6, IL-1,
IL-8, ICAM-1, VCAM-1, and VEGF. Such diseases or disorders may, in
some embodiments, involve undesirable proliferation of certain
cells, as in the case of cancer (e.g., solid tumors, leukemias,
myelomas, lymphomas, and other cancers), fibrosis associated with
organ failure, or excessive scarring. Non limiting examples of the
disease or disorder include: lupus, rheumatoid arthritis,
juvenile-onset diabetes, multiple sclerosis, psoriasis, and Crohn's
disease. Further non-limiting examples include cardiovascular
diseases, such as atherosclerosis, heart failure, myocardial
infarction, acute coronary syndrome, restenosis following vascular
surgery, hypertension, and vasculitis; neurodegenerative or
neuromuscular diseases such as Alzheimer's disease, Parkinson's
disease, Huntington's disease, ALS, and muscular dystrophy;
neurological disorders such as epilepsy and dystonia;
neuropsychiatric conditions such as major depression, bipolar
disorder, post-traumatic stress disorder, schizophrenia, anorexia
nervosa, ADHD, and autism-spectrum disorders; retinal diseases such
as macular degeneration, diabetic retinopathy, glaucoma, and
retinitis; chronic and acute pain syndromes, including inflammatory
and neuropathic pain; hearing loss and tinnitus; diabetes and
complications of diabetes, including metabolic syndrome, diabetic
nephropathy, diabetic neuropathy and diabetic ulcers; respiratory
diseases such as asthma, chronic obstructive pulmonary disease,
acute respiratory distress syndrome, and cystic fibrosis;
inflammatory bowel diseases; osteoporosis, osteoarthritis, and
other degenerative conditions of bone and cartilage; acute or
chronic organ failure, including renal failure, liver failure
(including cirrhosis and hepatitis), and pancreatitis;
ischemia-reperfusion injury associated with thrombotic or
hemorrhagic stroke, subarachnoid hemorrhage, cerebral vasospasm,
myocardial infarction, shock, or trauma; complications of organ or
tissue transplantation including acute or chronic transplant
failure or rejection and graft-versus-host disease; skin diseases
including atopic dermatitis and acne; sepsis and septic shock;
excessive inflammation associated with infection, including
respiratory inflammation associated with influenza and upper
respiratory infections; mucositis associated with cancer therapy,
including radiation therapy or chemotherapy; and severe burns.
Methods of synthesizing compounds of the present disclosure are
also contemplated. In particular embodiments, such methods can
comprise a method of making a target compound defined of the
formula:
##STR00028## wherein R.sub.a is alkoxy.sub.(C1-4), comprising
reacting a compound of the formula:
##STR00029## with an oxidizing agent under a set of conditions to
form the target compound.
Kits are also contemplated by the present disclosure, such as a kit
comprising: a compound of the present disclosure; and instructions
which comprise one or more forms of information selected from the
group consisting of indicating a disease state for which the
compound is to be administered, storage information for the
compound, dosing information and instructions regarding how to
administer the compound. The kit may comprise a compound of the
present disclosure in a multiple dose form.
Other objects, features and advantages of the present disclosure
will become apparent from the following detailed description. It
should be understood, however, that the detailed description and
the specific examples, while indicating specific embodiments of the
invention, are given by way of illustration only, since various
changes and modifications within the spirit and scope of the
invention will become apparent to those skilled in the art from
this detailed description. Note that simply because a particular
compound is ascribed to one particular generic formula doesn't mean
that it cannot also belong to another generic formula.
BRIEF DESCRIPTION OF THE DRAWINGS
The following drawings form part of the present specification and
are included to further demonstrate certain aspects of the present
disclosure. The invention may be better understood by reference to
one of these drawings in combination with the detailed description
of specific embodiments presented herein.
FIGS. 1-8 and 32-34. Inhibition of NO Production.
RAW264.7 macrophages were pre-treated with DMSO or drugs at various
concentrations (nM) for 2 hours, then treated with 20 ng/ml
IFN.gamma. for 24 hours. NO concentration in media was determined
using a Griess reagent system; cell viability was determined using
WST-1 reagent.
FIG. 9. Suppression of COX-2 Induction.
RAW264.7 cells were pre-treated for 2 hours with indicated
compounds and subsequently stimulated with 10 ng/ml IFN.gamma. for
an additional 24 hours. COX-2 protein levels were assayed by
immunoblotting. Actin was used as a loading control. RTA 402 and
RTA 404 refer to comparison compounds 402 and 404 (see Example
1).
FIGS. 10-12. Inhibition of IL-6 Induced STAT3 Phosphorylation.
HeLa cells were treated with the indicated compounds and
concentrations for 6 hours and subsequently stimulated with 20
ng/ml IL-6 for 15 minutes. Phosphorylated STAT3 and total STAT3
levels were assayed by immunoblotting. Compounds 402-52 and 402-53
are comparison compounds (see Example 1).
FIG. 13. Suppression of IL-6 Induced STAT3 Phosphorylation.
HeLa cells were treated with DMSO or the indicated compounds at 2
.mu.M for 6 hours and subsequently stimulated with 20 ng/ml IL-6
for 15 minutes. Phosphorylated STAT3 and total STAT3 levels were
assayed by immunoblotting. Compounds 402-54, 402-55 and 402-56 are
comparison compounds (see Example 1).
FIG. 14. Inhibition of TNF.alpha.-induced I.kappa.B.alpha.
degradation.
HeLa cells were treated with indicated compounds and concentrations
for 6 hours and subsequently stimulated with 20 ng/ml TNF.alpha.
for 15 minutes. Lysates were analyzed with antibodies against
I.kappa.B.alpha. and actin.
FIGS. 15 and 16. Inhibition of NF.kappa.B Activation.
HeLa cells were transfected with pNF-.kappa.B-Luc (inducible) and
pRL-TK (constitutive) reporter plasmids. Twenty-four hours later
cells were pre-treated with the indicated compounds for 2 hours.
DMSO served as a vehicle control. Following pre-treatment, cells
were stimulated with 20 ng/ml TNF.alpha. for 3 hours. Reporter
activity was measured using DualGlo luciferase reporter assay and
pNF-.kappa.B luciferase activity was normalized against pRL-TK
luciferase activity. Fold-induction of mean luciferase activity
relative to unstimulated (-TNF.alpha.) samples is shown. Error bars
represent the SD of the mean of 6 samples.
FIGS. 17-20. Induction of HO-1.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours. HO-1 mRNA
levels were quantified using qPCR and were normalized relative to a
DMSO-treated sample run in parallel. Values are averages of
duplicate wells.
FIG. 21. Induction of HO-1, TrxR1 and .gamma.-GCS.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours. HO-1,
thioredoxin reductase-1 (TrxR1), and .gamma.-glutamylcysteine
synthetase (.gamma.-GCS) mRNA levels were quantified using qPCR and
were normalized relative to a DMSO-treated sample run in parallel.
Values are averages of duplicate wells.
FIG. 22. Induction of TrxR1.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours.
Thioredoxin reductase-1 (TrxR1) mRNA levels were quantified using
qPCR and were normalized relative to a DMSO-treated sample run in
parallel. Values are averages of duplicate wells. Compounds 401,
402-19 and 402-53 are comparison compounds (see Example 1).
Comparison with the results of FIG. 25 demonstrates that higher
concentrations of 402-02 and 404-02 are required to approach
effects seen with the unsaturated counterpart compounds 402 and
404.
FIG. 23. Induction of .gamma.-GCS.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours.
.gamma.-glutamylcysteine synthetase (.gamma.-GCS) mRNA levels were
quantified using qPCR and were normalized relative to a
DMSO-treated sample run in parallel. Values are averages of
duplicate wells. Comparison with the results of FIG. 26
demonstrates that higher concentrations of 402-02 and 404-02 are
required to approach effects seen with the unsaturated counterpart
compounds 402 and 404.
FIG. 24. Induction of Ferritin Heavy Chain.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours. Ferritin
heavy chain mRNA levels were quantified using qPCR and were
normalized relative to a DMSO-treated sample run in parallel.
Values are averages of duplicate wells. Comparison with the results
of FIG. 27 demonstrates that higher concentrations of 402-02 and
404-02 are required to approach effects seen with the unsaturated
counterpart compounds 402 and 404.
FIG. 25. Induction of TrxR1.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours.
Thioredoxin reductase-1 (TrxR1) mRNA levels were quantified using
qPCR and were normalized relative to a DMSO-treated sample run in
parallel. Values are averages of duplicate wells. Comparison with
the results of FIG. 22 demonstrates that higher concentrations of
402-02 and 404-02 are required to approach effects seen with the
unsaturated counterpart compounds 402 and 404.
FIG. 26. Induction of .gamma.-GCS.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours.
.gamma.-glutamylcysteine synthetase (.gamma.-GCS) mRNA levels were
quantified using qPCR and were normalized relative to a
DMSO-treated sample run in parallel. Values are averages of
duplicate wells. Comparison with the results of FIG. 23
demonstrates that higher concentrations of 402-02 and 404-02 are
required to approach effects seen with the unsaturated counterpart
compounds 402 and 404.
FIG. 27. Induction of Ferritin Heavy Chain.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours. Ferritin
heavy chain mRNA levels were quantified using qPCR and were
normalized relative to a DMSO-treated sample run in parallel.
Values are averages of duplicate wells. Comparison with the results
of FIG. 24 demonstrates that higher concentrations of 402-02 and
404-02 are required to approach effects seen with the unsaturated
counterpart compounds 402 and 404.
FIG. 28--CDDO-TFEA (TP-500) is Detected at Higher Levels in Mouse
Brain than CDDO-EA (TP-319).
CD-1 mice were fed either 200 or 400 mg/kg diet of either TP-319 or
TP-500 for 3.5 days, and TP levels in the brains of the mice were
analyzed by LC/MS. The structures of TP-319 and TP-500 are shown
below.
FIGS. 29 A & B--Weight Change Data from a Head to Head Tox
Study of Compounds 401 (FIG. 29A) Versus 401-02 (FIG. 29B).
Compounds were assessed for toxicity in mice in a 14-day study.
Each compound was formulated in sesame oil and administered daily
by oral gavage at doses of 10, 50, 100, or 250 mg/kg (n=4 per
group).
FIGS. 30 A & B--Weight Change Data from a Head to Head Tox
Study of Compounds 402 (FIG. 30A) Versus 402-02 (FIG. 30B).
Compounds were assessed for toxicity in mice in a 14-day study.
Each compound was formulated in sesame oil and administered daily
by oral gavage at doses of 10, 50, 100, or 250 mg/kg (n=4 per
group).
FIGS. 31A & B--Weight Change Data from a Head to Head Tox Study
of Compounds 404 (FIG. 31A) Versus 404-02 (FIG. 31B).
Compounds were assessed for toxicity in mice in a 14-day study.
Each compound was formulated in sesame oil and administered daily
by oral gavage at doses of 10, 50, 100, or 250 mg/kg (n=4 per
group).
DESCRIPTION OF ILLUSTRATIVE EMBODIMENTS
Disclosed herein are, for example, new compounds with antioxidant
and anti-inflammatory properties, methods for their manufacture,
and methods for their use, including for the treatment and/or
prevention of disease.
I. DEFINITIONS
As used herein, "hydrogen" means --H; "hydroxy" means --OH; "oxo"
means .dbd.O; "halo" means independently --F, --Cl, --Br or --I;
"amino" means --NH.sub.2 (see below for definitions of groups
containing the term amino, e.g., alkylamino); "hydroxyamino" means
--NHOH; "nitro" means --NO.sub.2; imino means .dbd.NH (see below
for definitions of groups containing the term imino, e.g.,
alkylamino); "cyano" means --CN; "azido" means --N.sub.3;
"mercapto" means --SH; "thio" means .dbd.S; "sulfonamido" means
--NHS(O).sub.2-- (see below for definitions of groups containing
the term sulfonamido, e.g., alkylsulfonamido); "sulfonyl" means
--S(O).sub.2-- (see below for definitions of groups containing the
term sulfonyl, e.g., alkylsulfonyl); and "silyl" means --SiH.sub.3
(see below for definitions of group(s) containing the term silyl,
e.g., alkylsilyl).
For the groups below, the following parenthetical subscripts
further define the groups as follows: "(Cn)" defines the exact
number (n) of carbon atoms in the group. "(C.ltoreq.n)" defines the
maximum number (n) of carbon atoms that can be in the group, with
the minimum number of carbon atoms in such at least one, but
otherwise as small as possible for the group in question. E.g., it
is understood that the minimum number of carbon atoms in the group
"alkenyl.sub.(C.ltoreq.8)" is 2. For example,
"alkoxy.sub.(C.ltoreq.10)" designates those alkoxy groups having
from 1 to 10 carbon atoms (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10,
or any range derivable therein (e.g., 3-10 carbon atoms)). (Cn-n')
defines both the minimum (n) and maximum number (n') of carbon
atoms in the group. Similarly, "alkyl.sub.(C2-10)" designates those
alkyl groups having from 2 to 10 carbon atoms (e.g., 2, 3, 4, 5, 6,
7, 8, 9, or 10, or any range derivable therein (e.g., 3-10 carbon
atoms)).
The term "alkyl" when used without the "substituted" modifier
refers to a non-aromatic monovalent group with a saturated carbon
atom as the point of attachment, a linear or branched, cyclo,
cyclic or acyclic structure, no carbon-carbon double or triple
bonds, and no atoms other than carbon and hydrogen. The groups,
--CH.sub.3 (Me), --CH.sub.2CH.sub.3 (Et),
--CH.sub.2CH.sub.2CH.sub.3 (n-Pr), --CH(CH.sub.3).sub.2 (iso-Pr),
--CH(CH.sub.2).sub.2 (cyclopropyl),
--CH.sub.2CH.sub.2CH.sub.2CH.sub.3 (n-Bu),
--CH(CH.sub.3)CH.sub.2CH.sub.3 (sec-butyl),
--CH.sub.2CH(CH.sub.3).sub.2 (iso-butyl), --C(CH.sub.3).sub.3
(tert-butyl), --CH.sub.2C(CH.sub.3).sub.3 (neo-pentyl), cyclobutyl,
cyclopentyl, cyclohexyl, and cyclohexylmethyl are non-limiting
examples of alkyl groups. The term "substituted alkyl" refers to a
non-aromatic monovalent group with a saturated carbon atom as the
point of attachment, a linear or branched, cyclo, cyclic or acyclic
structure, no carbon-carbon double or triple bonds, and at least
one atom independently selected from the group consisting of N, O,
F, Cl, Br, I, Si, P, and S. The following groups are non-limiting
examples of substituted alkyl groups: --CH.sub.2OH, --CH.sub.2Cl,
--CH.sub.2Br, --CH.sub.2SH, --CF.sub.3, --CH.sub.2CN,
--CH.sub.2C(O)H, --CH.sub.2C(O)OH, --CH.sub.2C(O)OCH.sub.3,
--CH.sub.2C(O)NH.sub.2, --CH.sub.2C(O)NHCH.sub.3,
--CH.sub.2C(O)CH.sub.3, --CH.sub.2OCH.sub.3,
--CH.sub.2OCH.sub.2CF.sub.3, --CH.sub.2OC(O)CH.sub.3,
--CH.sub.2NH.sub.2, --CH.sub.2NHCH.sub.3,
--CH.sub.2N(CH.sub.3).sub.2, --CH.sub.2CH.sub.2Cl,
--CH.sub.2CH.sub.2OH, --CH.sub.2CF.sub.3,
--CH.sub.2CH.sub.2OC(O)CH.sub.3,
--CH.sub.2CH.sub.2NHCO.sub.2C(CH.sub.3).sub.3, and
--CH.sub.2Si(CH.sub.3).sub.3.
The term "alkanediyl" when used without the "substituted" modifier
refers to a non-aromatic divalent group, wherein the alkanediyl
group is attached with two .sigma.-bonds, with one or two saturated
carbon atom(s) as the point(s) of attachment, a linear or branched,
cyclo, cyclic or acyclic structure, no carbon-carbon double or
triple bonds, and no atoms other than carbon and hydrogen. The
groups, --CH.sub.2-- (methylene), --CH.sub.2CH.sub.2--,
CH.sub.2C(CH.sub.3).sub.2CH.sub.2--, --CH.sub.2CH.sub.2CH.sub.2--,
and
##STR00030## are non-limiting examples of alkanediyl groups. The
term "substituted alkanediyl" refers to a non-aromatic monovalent
group, wherein the alkynediyl group is attached with two
.sigma.-bonds, with one or two saturated carbon atom(s) as the
point(s) of attachment, a linear or branched, cyclo, cyclic or
acyclic structure, no carbon-carbon double or triple bonds, and at
least one atom independently selected from the group consisting of
N, O, F, Cl, Br, I, Si, P, and S. The following groups are
non-limiting examples of substituted alkanediyl groups: --CH(F)--,
--CF.sub.2--, --CH(Cl)--, --CH(OH)--, --CH(OCH.sub.3)--, and
--CH.sub.2CH(Cl)--.
The term "alkenyl" when used without the "substituted" modifier
refers to a monovalent group with a nonaromatic carbon atom as the
point of attachment, a linear or branched, cyclo, cyclic or acyclic
structure, at least one nonaromatic carbon-carbon double bond, no
carbon-carbon triple bonds, and no atoms other than carbon and
hydrogen. Non-limiting examples of alkenyl groups include:
--CH.dbd.CH.sub.2 (vinyl), --CH.dbd.CHCH.sub.3,
--CH.dbd.CHCH.sub.2CH.sub.3, --CH.sub.2CH.dbd.CH.sub.2 (allyl),
--CH.sub.2CH.dbd.CHCH.sub.3, and --CH.dbd.CH--C.sub.6H.sub.5. The
term "substituted alkenyl" refers to a monovalent group with a
nonaromatic carbon atom as the point of attachment, at least one
nonaromatic carbon-carbon double bond, no carbon-carbon triple
bonds, a linear or branched, cyclo, cyclic or acyclic structure,
and at least one atom independently selected from the group
consisting of N, O, F, Cl, Br, I, Si, P, and S. The groups,
--CH.dbd.CHF, --CH.dbd.CHCl and --CH.dbd.CHBr, are non-limiting
examples of substituted alkenyl groups.
The term "alkenediyl" when used without the "substituted" modifier
refers to a non-aromatic divalent group, wherein the alkenediyl
group is attached with two .sigma.-bonds, with two carbon atoms as
points of attachment, a linear or branched, cyclo, cyclic or
acyclic structure, at least one nonaromatic carbon-carbon double
bond, no carbon-carbon triple bonds, and no atoms other than carbon
and hydrogen. The groups, --CH.dbd.CH--,
--CH.dbd.C(CH.sub.3)CH.sub.2--, --CH.dbd.CHCH.sub.2--, and
##STR00031## are non-limiting examples of alkenediyl groups. The
term "substituted alkenediyl" refers to a non-aromatic divalent
group, wherein the alkenediyl group is attached with two
.sigma.-bonds, with two carbon atoms as points of attachment, a
linear or branched, cyclo, cyclic or acyclic structure, at least
one nonaromatic carbon-carbon double bond, no carbon-carbon triple
bonds, and at least one atom independently selected from the group
consisting of N, O, F, Cl, Br, I, Si, P, and S. The following
groups are non-limiting examples of substituted alkenediyl groups:
--CF.dbd.CH--, --C(OH).dbd.CH--, and --CH.sub.2CH.dbd.C(Cl)--.
The term "alkynyl" when used without the "substituted" modifier
refers to a monovalent group with a nonaromatic carbon atom as the
point of attachment, a linear or branched, cyclo, cyclic or acyclic
structure, at least one carbon-carbon triple bond, and no atoms
other than carbon and hydrogen. The groups, --C.ident.CH,
--C.ident.CCH.sub.3, --C.ident.CC.sub.6H.sub.5 and
--CH.sub.2C.ident.CCH.sub.3, are non-limiting examples of alkynyl
groups. The term "substituted alkynyl" refers to a monovalent group
with a nonaromatic carbon atom as the point of attachment and at
least one carbon-carbon triple bond, a linear or branched, cyclo,
cyclic or acyclic structure, and at least one atom independently
selected from the group consisting of N, O, F, Cl, Br, I, Si, P,
and S. The group, --C.ident.CSi(CH.sub.3).sub.3, is a non-limiting
example of a substituted alkynyl group.
The term "alkynediyl" when used without the "substituted" modifier
refers to a non-aromatic divalent group, wherein the alkynediyl
group is attached with two .sigma.-bonds, with two carbon atoms as
points of attachment, a linear or branched, cyclo, cyclic or
acyclic structure, at least one carbon-carbon triple bond, and no
atoms other than carbon and hydrogen. The groups, --C.ident.C--,
--C.ident.CCH.sub.2--, and --C.ident.CCH(CH.sub.3)-- are
non-limiting examples of alkynediyl groups. The term "substituted
alkynediyl" refers to a non-aromatic divalent group, wherein the
alkynediyl group is attached with two .sigma.-bonds, with two
carbon atoms as points of attachment, a linear or branched, cyclo,
cyclic or acyclic structure, at least one carbon-carbon triple
bond, and at least one atom independently selected from the group
consisting of N, O, F, Cl, Br, I, Si, P, and S. The groups
--C.ident.CCFH-- and --C.ident.CHCH(Cl)-- are non-limiting examples
of substituted alkynediyl groups.
The term "aryl" when used without the "substituted" modifier refers
to a monovalent group with an aromatic carbon atom as the point of
attachment, said carbon atom forming part of a six-membered
aromatic ring structure wherein the ring atoms are all carbon, and
wherein the monovalent group consists of no atoms other than carbon
and hydrogen. Non-limiting examples of aryl groups include phenyl
(Ph), methylphenyl, (dimethyl)phenyl,
--C.sub.6H.sub.4CH.sub.2CH.sub.3 (ethylphenyl),
--C.sub.6H.sub.4CH.sub.2CH.sub.2CH.sub.3 (propylphenyl),
--C.sub.6H.sub.4CH(CH.sub.3).sub.2,
--C.sub.6H.sub.4CH(CH.sub.2).sub.2,
--C.sub.6H.sub.3(CH.sub.3)CH.sub.2CH.sub.3 (methylethylphenyl),
--C.sub.6H.sub.4CH.dbd.CH.sub.2 (vinylphenyl),
--C.sub.6H.sub.4CH.dbd.CHCH.sub.3, --C.sub.6H.sub.4C.ident.CH,
--C.sub.6H.sub.4C.ident.CCH.sub.3, naphthyl, and the monovalent
group derived from biphenyl. The term "substituted aryl" refers to
a monovalent group with an aromatic carbon atom as the point of
attachment, said carbon atom forming part of a six-membered
aromatic ring structure wherein the ring atoms are all carbon, and
wherein the monovalent group further has at least one atom
independently selected from the group consisting of N, O, F, Cl,
Br, I, Si, P, and S. Non-limiting examples of substituted aryl
groups include the groups: --C.sub.6H.sub.4F, --C.sub.6H.sub.4Cl,
--C.sub.6H.sub.4Br, --C.sub.6H.sub.4I, --C.sub.6H.sub.4OH,
--C.sub.6H.sub.4OCH.sub.3, --C.sub.6H.sub.4OCH.sub.2CH.sub.3,
--C.sub.6H.sub.4OC(O)CH.sub.3, C.sub.6H.sub.4NH.sub.2,
C.sub.6H.sub.4NHCH.sub.3, C.sub.6H.sub.4N(CH.sub.3).sub.2,
C.sub.6H.sub.4CH.sub.2OH, C.sub.6H.sub.4CH.sub.2OC(O)CH.sub.3,
C.sub.6H.sub.4CH.sub.2NH.sub.2, C.sub.6H.sub.4CF.sub.3,
--C.sub.6H.sub.4CN, --C.sub.6H.sub.4CHO, --C.sub.6H.sub.4CHO,
--C.sub.6H.sub.4C(O)CH.sub.3, --C.sub.6H.sub.4C(O)C.sub.6H.sub.5,
--C.sub.6H.sub.4CO.sub.2H, --C.sub.6H.sub.4CO.sub.2CH.sub.3,
--C.sub.6H.sub.4CONH.sub.2, --C.sub.6H.sub.4CONHCH.sub.3, and
--C.sub.6H.sub.4CON(CH.sub.3).sub.2.
The term "arenediyl" when used without the "substituted" modifier
refers to a divalent group, wherein the arenediyl group is attached
with two .sigma.-bonds, with two aromatic carbon atoms as points of
attachment, said carbon atoms forming part of one or more
six-membered aromatic ring structure(s) wherein the ring atoms are
all carbon, and wherein the monovalent group consists of no atoms
other than carbon and hydrogen. Non-limiting examples of arenediyl
groups include:
##STR00032## The term "substituted arenediyl" refers to a divalent
group, wherein the arenediyl group is attached with two
.sigma.-bonds, with two aromatic carbon atoms as points of
attachment, said carbon atoms forming part of one or more
six-membered aromatic rings structure(s), wherein the ring atoms
are all carbon, and wherein the divalent group further has at least
one atom independently selected from the group consisting of N, O,
F, Cl, Br, I, Si, P, and S.
The term "aralkyl" when used without the "substituted" modifier
refers to the monovalent group -alkanediyl-aryl, in which the terms
alkanediyl and aryl are each used in a manner consistent with the
definitions provided above. Non-limiting examples of aralkyls are:
phenylmethyl (benzyl, Bn), 1-phenyl-ethyl, 2-phenyl-ethyl, indenyl
and 2,3-dihydro-indenyl, provided that indenyl and
2,3-dihydro-indenyl are only examples of aralkyl in so far as the
point of attachment in each case is one of the saturated carbon
atoms. When the term "aralkyl" is used with the "substituted"
modifier, either one or both the alkanediyl and the aryl is
substituted. Non-limiting examples of substituted aralkyls are:
(3-chlorophenyl)-methyl, 2-oxo-2-phenyl-ethyl
(phenylcarbonylmethyl), 2-chloro-2-phenyl-ethyl, chromanyl where
the point of attachment is one of the saturated carbon atoms, and
tetrahydroquinolinyl where the point of attachment is one of the
saturated atoms.
The term "heteroaryl" when used without the "substituted" modifier
refers to a monovalent group with an aromatic carbon atom or
nitrogen atom as the point of attachment, said carbon atom or
nitrogen atom forming part of an aromatic ring structure wherein at
least one of the ring atoms is nitrogen, oxygen or sulfur, and
wherein the monovalent group consists of no atoms other than
carbon, hydrogen, aromatic nitrogen, aromatic oxygen and aromatic
sulfur. Non-limiting examples of aryl groups include acridinyl,
furanyl, imidazoimidazolyl, imidazopyrazolyl, imidazopyridinyl,
imidazopyrimidinyl, indolyl, indazolinyl, methylpyridyl, oxazolyl,
phenylimidazolyl, pyridyl, pyrrolyl, pyrimidyl, pyrazinyl,
quinolyl, quinazolyl, quinoxalinyl, tetrahydroquinolinyl, thienyl,
triazinyl, pyrrolopyridinyl, pyrrolopyrimidinyl, pyrrolopyrazinyl,
pyrrolotriazinyl, pyrroloimidazolyl, chromenyl (where the point of
attachment is one of the aromatic atoms), and chromanyl (where the
point of attachment is one of the aromatic atoms). The term
"substituted heteroaryl" refers to a monovalent group with an
aromatic carbon atom or nitrogen atom as the point of attachment,
said carbon atom or nitrogen atom forming part of an aromatic ring
structure wherein at least one of the ring atoms is nitrogen,
oxygen or sulfur, and wherein the monovalent group further has at
least one atom independently selected from the group consisting of
non-aromatic nitrogen, non-aromatic oxygen, non aromatic sulfur F,
Cl, Br, I, Si, and P.
The term "heteroarenediyl" when used without the "substituted"
modifier refers to a divalent group, wherein the heteroarenediyl
group is attached with two .sigma.-bonds, with an aromatic carbon
atom or nitrogen atom as the point of attachment, said carbon atom
or nitrogen atom two aromatic atoms as points of attachment, said
carbon atoms forming part of one or more six-membered aromatic ring
structure(s) wherein the ring atoms are all carbon, and wherein the
monovalent group consists of no atoms other than carbon and
hydrogen. Non-limiting examples of heteroarenediyl groups
include:
##STR00033## The term "substituted heteroarenediyl" refers to a
divalent group, wherein the heteroarenediyl group is attached with
two .sigma.-bonds, with two aromatic carbon atoms as points of
attachment, said carbon atoms forming part of one or more
six-membered aromatic rings structure(s), wherein the ring atoms
are all carbon, and wherein the divalent group further has at least
one atom independently selected from the group consisting of N, O,
F, Cl, Br, I, Si, P, and S.
The term "heteroaralkyl" when used without the "substituted"
modifier refers to the monovalent group -alkanediyl-heteroaryl, in
which the terms alkanediyl and heteroaryl are each used in a manner
consistent with the definitions provided above. Non-limiting
examples of aralkyls are: pyridylmethyl, and thienylmethyl. When
the term "heteroaralkyl" is used with the "substituted" modifier,
either one or both the alkanediyl and the heteroaryl is
substituted.
The term "acyl" when used without the "substituted" modifier refers
to a monovalent group with a carbon atom of a carbonyl group as the
point of attachment, further having a linear or branched, cyclo,
cyclic or acyclic structure, further having no additional atoms
that are not carbon or hydrogen, beyond the oxygen atom of the
carbonyl group. The groups, --CHO, --C(O)CH.sub.3 (acetyl, Ac),
--C(O)CH.sub.2CH.sub.3, --C(O)CH.sub.2CH.sub.2CH.sub.3,
--C(O)CH(CH.sub.3).sub.2, --C(O)CH(CH.sub.2).sub.2,
--C(O)C.sub.6H.sub.5, --C(O)C.sub.6H.sub.4CH.sub.3,
--C(O)C.sub.6H.sub.4CH.sub.2CH.sub.3,
COC.sub.6H.sub.3(CH.sub.3).sub.2, and --C(O)CH.sub.2C.sub.6H.sub.5,
are non-limiting examples of acyl groups. The term "acyl" therefore
encompasses, but is not limited to groups sometimes referred to as
"alkyl carbonyl" and "aryl carbonyl" groups. The term "substituted
acyl" refers to a monovalent group with a carbon atom of a carbonyl
group as the point of attachment, further having a linear or
branched, cyclo, cyclic or acyclic structure, further having at
least one atom, in addition to the oxygen of the carbonyl group,
independently selected from the group consisting of N, O, F, Cl,
Br, I, Si, P, and S. The groups, --C(O)CH.sub.2CF.sub.3,
--CO.sub.2H (carboxyl), --CO.sub.2CH.sub.3 (methylcarboxyl),
--CO.sub.2CH.sub.2CH.sub.3, --CO.sub.2CH.sub.2CH.sub.2CH.sub.3,
CO.sub.2C.sub.6H.sub.5, --CO.sub.2CH(CH.sub.3).sub.2,
--CO.sub.2CH(CH.sub.2).sub.2, --C(O)NH.sub.2 (carbamoyl),
--C(O)NHCH.sub.3, --C(O)NHCH.sub.2CH.sub.3,
--CONHCH(CH.sub.3).sub.2, --CONHCH(CH.sub.2).sub.2,
--CON(CH.sub.3).sub.2, --CONHCH.sub.2CF.sub.3, --CO-pyridyl,
--CO-imidazoyl, and --C(O)N.sub.3, are non-limiting examples of
substituted acyl groups. The term "substituted acyl" encompasses,
but is not limited to, "heteroaryl carbonyl" groups.
The term "alkylidene" when used without the "substituted" modifier
refers to the divalent group .dbd.CRR', wherein the alkylidene
group is attached with one .sigma.-bond and one .pi.-bond, in which
R and R' are independently hydrogen, alkyl, or R and R' are taken
together to represent alkanediyl. Non-limiting examples of
alkylidene groups include: .dbd.CH.sub.2,
.dbd.CH(CH.sub.2CH.sub.3), and .dbd.C(CH.sub.3).sub.2. The term
"substituted alkylidene" refers to the group .dbd.CRR', wherein the
alkylidene group is attached with one .sigma.-bond and one
.pi.-bond, in which R and R' are independently hydrogen, alkyl,
substituted alkyl, or R and R' are taken together to represent a
substituted alkanediyl, provided that either one of R and R' is a
substituted alkyl or R and R' are taken together to represent a
substituted alkanediyl.
The term "alkoxy" when used without the "substituted" modifier
refers to the group --OR, in which R is an alkyl, as that term is
defined above. Non-limiting examples of alkoxy groups include:
--OCH.sub.3, --OCH.sub.2CH.sub.3, --OCH.sub.2CH.sub.2CH.sub.3,
--OCH(CH.sub.3).sub.2, --OCH(CH.sub.2).sub.2, --O-cyclopentyl, and
--O-cyclohexyl. The term "substituted alkoxy" refers to the group
--OR, in which R is a substituted alkyl, as that term is defined
above. For example, --OCH.sub.2CF.sub.3 is a substituted alkoxy
group.
Similarly, the terms "alkenyloxy", "alkynyloxy", "aryloxy",
"aralkoxy", "heteroaryloxy", "heteroaralkoxy" and "acyloxy", when
used without the "substituted" modifier, refers to groups, defined
as --OR, in which R is alkenyl, alkynyl, aryl, aralkyl, heteroaryl,
heteroaralkyl and acyl, respectively, as those terms are defined
above. When any of the terms alkenyloxy, alkynyloxy, aryloxy,
aralkyloxy and acyloxy is modified by "substituted," it refers to
the group --OR, in which R is substituted alkenyl, alkynyl, aryl,
aralkyl, heteroaryl, heteroaralkyl and acyl, respectively.
The term "alkylamino" when used without the "substituted" modifier
refers to the group --NHR, in which R is an alkyl, as that term is
defined above. Non-limiting examples of alkylamino groups include:
--NHCH.sub.3, --NHCH.sub.2CH.sub.3, --NHCH.sub.2CH.sub.2CH.sub.3,
--NHCH(CH.sub.3).sub.2, --NHCH(CH.sub.2).sub.2,
--NHCH.sub.2CH.sub.2CH.sub.2CH.sub.3,
--NHCH(CH.sub.3)CH.sub.2CH.sub.3, --NHCH.sub.2CH(CH.sub.3).sub.2,
--NHC(CH.sub.3).sub.3, --NH-cyclopentyl, and --NH-cyclohexyl. The
term "substituted alkylamino" refers to the group --NHR, in which R
is a substituted alkyl, as that term is defined above. For example,
--NHCH.sub.2CF.sub.3 is a substituted alkylamino group.
The term "dialkylamino" when used without the "substituted"
modifier refers to the group --NRR', in which R and R' can be the
same or different alkyl groups, or R and R' can be taken together
to represent an alkanediyl having two or more saturated carbon
atoms, at least two of which are attached to the nitrogen atom.
Non-limiting examples of dialkylamino groups include:
--NHC(CH.sub.3).sub.3, --N(CH.sub.3)CH.sub.2CH.sub.3,
--N(CH.sub.2CH.sub.3).sub.2, N-pyrrolidinyl, and N-piperidinyl. The
term "substituted dialkylamino" refers to the group --NRR', in
which R and R' can be the same or different substituted alkyl
groups, one of R or R' is an alkyl and the other is a substituted
alkyl, or R and R' can be taken together to represent a substituted
alkanediyl with two or more saturated carbon atoms, at least two of
which are attached to the nitrogen atom.
The terms "alkoxyamino", "alkenylamino", "alkynylamino",
"arylamino", "aralkylamino", "heteroarylamino",
"heteroaralkylamino", and "alkylsulfonylamino" when used without
the "substituted" modifier, refers to groups, defined as --NHR, in
which R is alkoxy, alkenyl, alkynyl, aryl, aralkyl, heteroaryl,
heteroaralkyl and alkylsulfonyl, respectively, as those terms are
defined above. A non-limiting example of an arylamino group is
--NHC.sub.6H.sub.5. When any of the terms alkoxyamino,
alkenylamino, alkynylamino, arylamino, aralkylamino,
heteroarylamino, hetero aralkyl amino and alkylsulfonylamino is
modified by "substituted," it refers to the group --NHR, in which R
is substituted alkoxy, alkenyl, alkynyl, aryl, aralkyl, heteroaryl,
heteroaralkyl and alkylsulfonyl, respectively.
The term "amido" (acylamino), when used without the "substituted"
modifier, refers to the group --NHR, in which R is acyl, as that
term is defined above. A non-limiting example of an acylamino group
is --NHC(O)CH.sub.3. When the term amido is used with the
"substituted" modifier, it refers to groups, defined as --NHR, in
which R is substituted acyl, as that term is defined above. The
groups --NHC(O)OCH.sub.3 and --NHC(O)NHCH.sub.3 are non-limiting
examples of substituted amido groups.
The term "alkylimino" when used without the "substituted" modifier
refers to the group .dbd.NR, wherein the alkylimino group is
attached with one .sigma.-bond and one .pi.-bond, in which R is an
alkyl, as that term is defined above. Non-limiting examples of
alkylimino groups include: .dbd.NCH.sub.3, .dbd.NCH.sub.2CH.sub.3
and .dbd.N-cyclohexyl. The term "substituted alkylimino" refers to
the group .dbd.NR, wherein the alkylimino group is attached with
one .sigma.-bond and one .pi.-bond, in which R is a substituted
alkyl, as that term is defined above. For example,
.dbd.NCH.sub.2CF.sub.3 is a substituted alkylimino group.
Similarly, the terms "alkenylimino", "alkynylimino", "arylimino",
"aralkylimino", "heteroarylimino", "heteroaralkylimino" and
"acylimino", when used without the "substituted" modifier, refers
to groups, defined as .dbd.NR, wherein the alkylimino group is
attached with one .sigma.-bond and one .pi.-bond, in which R is
alkenyl, alkynyl, aryl, aralkyl, heteroaryl, heteroaralkyl and
acyl, respectively, as those terms are defined above. When any of
the terms alkenylimino, alkynylimino, arylimino, aralkylimino and
acylimino is modified by "substituted," it refers to the group
.dbd.NR, wherein the alkylimino group is attached with one
.sigma.-bond and one .pi.-bond, in which R is substituted alkenyl,
alkynyl, aryl, aralkyl, heteroaryl, heteroaralkyl and acyl,
respectively.
The term "fluoroalkyl" when used without the "substituted" modifier
refers to an alkyl, as that term is defined above, in which one or
more fluorines have been substituted for hydrogens. The groups,
--CH.sub.2F, --CF.sub.3, and --CH.sub.2CF.sub.3 are non-limiting
examples of fluoroalkyl groups. The term "substituted fluoroalkyl"
refers to a non-aromatic monovalent group with a saturated carbon
atom as the point of attachment, a linear or branched, cyclo,
cyclic or acyclic structure, at least one fluorine atom, no
carbon-carbon double or triple bonds, and at least one atom
independently selected from the group consisting of N, O, Cl, Br,
I, Si, P, and S. The following group is a non-limiting example of a
substituted fluoroalkyl: --CFHOH.
The term "alkylthio" when used without the "substituted" modifier
refers to the group --SR, in which R is an alkyl, as that term is
defined above. Non-limiting examples of alkylthio groups include:
--SCH.sub.3, --SCH.sub.2CH.sub.3, --SCH.sub.2CH.sub.2CH.sub.3,
--SCH(CH.sub.3).sub.2, --SCH(CH.sub.2).sub.2, --S-cyclopentyl, and
--S-cyclohexyl. The term "substituted alkylthio" refers to the
group --SR, in which R is a substituted alkyl, as that term is
defined above. For example, --SCH.sub.2CF.sub.3 is a substituted
alkylthio group.
Similarly, the terms "alkenylthio", "alkynylthio", "arylthio",
"aralkylthio", "heteroarylthio", "heteroaralkylthio", and
"acylthio", when used without the "substituted" modifier, refers to
groups, defined as --SR, in which R is alkenyl, alkynyl, aryl,
aralkyl, heteroaryl, heteroaralkyl and acyl, respectively, as those
terms are defined above. When any of the terms alkenylthio,
alkynylthio, arylthio, aralkylthio, heteroarylthio,
heteroaralkylthio, and acylthio is modified by "substituted," it
refers to the group --SR, in which R is substituted alkenyl,
alkynyl, aryl, aralkyl, heteroaryl, heteroaralkyl and acyl,
respectively.
The term "thioacyl" when used without the "substituted" modifier
refers to a monovalent group with a carbon atom of a thiocarbonyl
group as the point of attachment, further having a linear or
branched, cyclo, cyclic or acyclic structure, further having no
additional atoms that are not carbon or hydrogen, beyond the sulfur
atom of the carbonyl group. The groups, --CHS, --C(S)CH.sub.3,
--C(S)CH.sub.2CH.sub.3, --C(S)CH.sub.2CH.sub.2CH.sub.3,
--C(S)CH(CH.sub.3).sub.2, --C(S)CH(CH.sub.2).sub.2,
--C(S)C.sub.6H.sub.5, --C(S)C.sub.6H.sub.4CH.sub.3,
--C(S)C.sub.6H.sub.4CH.sub.2CH.sub.3,
--C(S)C.sub.6H.sub.3(CH.sub.3).sub.2, and
--C(S)CH.sub.2C.sub.6H.sub.5, are non-limiting examples of thioacyl
groups. The term "thioacyl" therefore encompasses, but is not
limited to, groups sometimes referred to as "alkyl thiocarbonyl"
and "aryl thiocarbonyl" groups. The term "substituted thioacyl"
refers to a radical with a carbon atom as the point of attachment,
the carbon atom being part of a thiocarbonyl group, further having
a linear or branched, cyclo, cyclic or acyclic structure, further
having at least one atom, in addition to the sulfur atom of the
carbonyl group, independently selected from the group consisting of
N, O, F, Cl, Br, I, Si, P, and S. The groups,
--C(S)CH.sub.2CF.sub.3, --C(S)O.sub.2H, --C(S)OCH.sub.3,
--C(S)OCH.sub.2CH.sub.3, --C(S)OCH.sub.2CH.sub.2CH.sub.3,
--C(S)OC.sub.6H.sub.5, --C(S)OCH(CH.sub.3).sub.2,
--C(S)OCH(CH.sub.2).sub.2, --C(S)NH.sub.2, and --C(S)NHCH.sub.3,
are non-limiting examples of substituted thioacyl groups. The term
"substituted thioacyl" encompasses, but is not limited to,
"heteroaryl thiocarbonyl" groups.
The term "alkylsulfonyl" when used without the "substituted"
modifier refers to the group --S(O).sub.2R, in which R is an alkyl,
as that term is defined above. Non-limiting examples of
alkylsulfonyl groups include: --S(O).sub.2CH.sub.3,
--S(O).sub.2CH.sub.2CH.sub.3, --S(O).sub.2CH.sub.2CH.sub.2CH.sub.3,
--S(O).sub.2CH(CH.sub.3).sub.2, --S(O).sub.2CH(CH.sub.2).sub.2,
--S(O).sub.2-cyclopentyl, and --S(O).sub.2-cyclohexyl. The term
"substituted alkylsulfonyl" refers to the group --S(O).sub.2R, in
which R is a substituted alkyl, as that term is defined above. For
example, --S(O).sub.2CH.sub.2CF.sub.3 is a substituted
alkylsulfonyl group.
Similarly, the terms "alkenylsulfonyl", "alkynylsulfonyl",
"arylsulfonyl", "aralkylsulfonyl", "heteroarylsulfonyl", and
"heteroaralkylsulfonyl" when used without the "substituted"
modifier, refers to groups, defined as --S(O).sub.2R, in which R is
alkenyl, alkynyl, aryl, aralkyl, heteroaryl, and heteroaralkyl,
respectively, as those terms are defined above. When any of the
terms alkenylsulfonyl, alkynylsulfonyl, arylsulfonyl,
aralkylsulfonyl, heteroarylsulfonyl, and heteroaralkylsulfonyl is
modified by "substituted," it refers to the group --S(O).sub.2R, in
which R is substituted alkenyl, alkynyl, aryl, aralkyl, heteroaryl
and heteroaralkyl, respectively.
The term "alkylammonium" when used without the "substituted"
modifier refers to a group, defined as --NH.sub.2R.sup.+,
--NHRR'.sup.+, or --NRR'R''.sup.+, in which R, R' and R'' are the
same or different alkyl groups, or any combination of two of R, R'
and R'' can be taken together to represent an alkanediyl.
Non-limiting examples of alkylammonium cation groups include:
--NH.sub.2(CH.sub.3).sup.+, --NH.sub.2(CH.sub.2CH.sub.3)+,
--NH.sub.2(CH.sub.2CH.sub.2CH.sub.3)+, --NH(CH.sub.3).sub.2.sup.+,
--NH(CH.sub.2CH.sub.3).sub.2.sup.+,
--NH(CH.sub.2CH.sub.2CH.sub.3).sub.2.sup.+,
--N(CH.sub.3).sub.3.sup.+,
--N(CH.sub.3)(CH.sub.2CH.sub.3).sub.2.sup.+,
--N(CH.sub.3).sub.2(CH.sub.2CH.sub.3).sup.+,
--NH.sub.2C(CH.sub.3).sub.3.sup.+, --NH(cyclopentyl).sub.2.sup.+,
and --NH.sub.2(cyclohexyl).sup.+. The term "substituted
alkylammonium" refers --NH.sub.2R.sup.+, --NHRR'.sup.+, or
--NRR'R''.sup.+, in which at least one of R, R' and R'' is a
substituted alkyl or two of R, R' and R'' can be taken together to
represent a substituted alkanediyl. When more than one of R, R' and
R'' is a substituted alkyl, they can be the same of different. Any
of R, R' and R'' that are not either substituted alkyl or
substituted alkanediyl, can be either alkyl, either the same or
different, or can be taken together to represent a alkanediyl with
two or more carbon atoms, at least two of which are attached to the
nitrogen atom shown in the formula.
The term "alkylsulfonium" when used without the "substituted"
modifier refers to the group --SRR'.sup.+, in which R and R' can be
the same or different alkyl groups, or R and R' can be taken
together to represent an alkanediyl. Non-limiting examples of
alkylsulfonium groups include: --SH(CH.sub.3).sup.+,
--SH(CH.sub.2CH.sub.3).sup.+, --SH(CH.sub.2CH.sub.2CH.sub.3).sup.+,
S(CH.sub.3).sub.2.sup.+, S(CH.sub.2CH.sub.3).sub.2.sup.+,
S(CH.sub.2CH.sub.2CH.sub.3).sub.2.sup.+, SH(cyclopentyl).sup.+, and
--SH(cyclohexyl).sup.+. The term "substituted alkylsulfonium"
refers to the group --SRR'.sup.+, in which R and R' can be the same
or different substituted alkyl groups, one of R or R' is an alkyl
and the other is a substituted alkyl, or R and R' can be taken
together to represent a substituted alkanediyl. For example,
--SH(CH.sub.2CF.sub.3).sup.+ is a substituted alkylsulfonium
group.
The term "alkylsilyl" when used without the "substituted" modifier
refers to a monovalent group, defined as --SiH.sub.2R, --SiHRR', or
--SiRR'R'', in which R, R' and R'' can be the same or different
alkyl groups, or any combination of two of R, R' and R'' can be
taken together to represent an alkanediyl. The groups,
--SiH.sub.2CH.sub.3, --SiH(CH.sub.3).sub.2, --Si(CH.sub.3).sub.3
and --Si(CH.sub.3).sub.2C(CH.sub.3).sub.3, are non-limiting
examples of unsubstituted alkylsilyl groups. The term "substituted
alkylsilyl" refers --SiH.sub.2R, --SiHRR', or --SiRR'R'', in which
at least one of R, R' and R'' is a substituted alkyl or two of R,
R' and R'' can be taken together to represent a substituted
alkanediyl. When more than one of R, R' and R'' is a substituted
alkyl, they can be the same of different. Any of R, R' and R'' that
are not either substituted alkyl or substituted alkanediyl, can be
either alkyl, either the same or different, or can be taken
together to represent a alkanediyl with two or more saturated
carbon atoms, at least two of which are attached to the silicon
atom.
In addition, atoms making up the compounds of the present
disclosure are intended to include all isotopic forms of such
atoms. Isotopes, as used herein, include those atoms having the
same atomic number but different mass numbers. By way of general
example and without limitation, isotopes of hydrogen include
tritium and deuterium, and isotopes of carbon include .sup.13C and
.sup.14C. Similarly, it is contemplated that one or more carbon
atom(s) of a compound of the present disclosure may be replaced by
a silicon atom(s). Furthermore, it is contemplated that one or more
oxygen atom(s) of a compound of the present disclosure may be
replaced by a sulfur or selenium atom(s).
A compound having a formula that is represented with a dashed bond
is intended to include the formulae optionally having zero, one or
more double bonds. Thus, for example, the structure
##STR00034## includes the structures
##STR00035## As will be understood by a person of skill in the art,
no one such ring atom forms part of more than one double bond.
Any undefined valency on an atom of a structure shown in this
application implicitly represents a hydrogen atom bonded to the
atom.
A ring structure shown with an unconnected "R" group, indicates
that any implicit hydrogen atom on that ring can be replaced with
that R group. In the case of a divalent R group (e.g., oxo, imino,
thio, alkylidene, etc.), any pair of implicit hydrogen atoms
attached to one atom of that ring can be replaced by that R group.
This concept is as exemplified below:
##STR00036## represents
##STR00037##
As used herein, a "chiral auxiliary" refers to a removable chiral
group that is capable of influencing the stereoselectivity of a
reaction. Persons of skill in the art are familiar with such
compounds, and many are commercially available.
The use of the word "a" or "an," when used in conjunction with the
term "comprising" in the claims and/or the specification may mean
"one," but it is also consistent with the meaning of "one or more,"
"at least one," and "one or more than one."
Throughout this application, the term "about" is used to indicate
that a value includes the inherent variation of error for the
device, the method being employed to determine the value, or the
variation that exists among the study subjects.
The terms "comprise," "have" and "include" are open-ended linking
verbs. Any forms or tenses of one or more of these verbs, such as
"comprises," "comprising," "has," "having," "includes" and
"including," are also open-ended. For example, any method that
"comprises," "has" or "includes" one or more steps is not limited
to possessing only those one or more steps and also covers other
unlisted steps.
The term "effective," as that term is used in the specification
and/or claims, means adequate to accomplish a desired, expected, or
intended result.
The term "hydrate" when used as a modifier to a compound means that
the compound has less than one (e.g., hemihydrate), one (e.g.,
monohydrate), or more than one (e.g., dihydrate) water molecules
associated with each compound molecule, such as in solid forms of
the compound.
As used herein, the term "IC.sub.50" refers to an inhibitory dose
which is 50% of the maximum response obtained.
An "isomer" of a first compound is a separate compound in which
each molecule contains the same constituent atoms as the first
compound, but where the configuration of those atoms in three
dimensions differs.
As used herein, the term "patient" or "subject" refers to a living
mammalian organism, such as a human, monkey, cow, sheep, goat, dog,
cat, mouse, rat, guinea pig, or transgenic species thereof. In
certain embodiments, the patient or subject is a primate.
Non-limiting examples of human subjects are adults, juveniles,
infants and fetuses.
"Pharmaceutically acceptable" means that which is useful in
preparing a pharmaceutical composition that is generally safe,
non-toxic and neither biologically nor otherwise undesirable and
includes that which is acceptable for veterinary use as well as
human pharmaceutical use.
"Pharmaceutically acceptable salts" means salts of compounds of the
present disclosure which are pharmaceutically acceptable, as
defined above, and which possess the desired pharmacological
activity. Such salts include acid addition salts formed with
inorganic acids such as hydrochloric acid, hydrobromic acid,
sulfuric acid, nitric acid, phosphoric acid, and the like; or with
organic acids such as 1,2-ethanedisulfonic acid,
2-hydroxyethanesulfonic acid, 2-naphthalenesulfonic acid,
3-phenylpropionic acid,
4,4'-methylenebis(3-hydroxy-2-ene-1-carboxylic acid),
4-methylbicyclo[2.2.2]oct-2-ene-1-carboxylic acid, acetic acid,
aliphatic mono- and dicarboxylic acids, aliphatic sulfuric acids,
aromatic sulfuric acids, benzenesulfonic acid, benzoic acid,
camphorsulfonic acid, carbonic acid, cinnamic acid, citric acid,
cyclopentanepropionic acid, ethanesulfonic acid, fumaric acid,
glucoheptonic acid, gluconic acid, glutamic acid, glycolic acid,
heptanoic acid, hexanoic acid, hydroxynaphthoic acid, lactic acid,
laurylsulfuric acid, maleic acid, malic acid, malonic acid,
mandelic acid, methanesulfonic acid, muconic acid,
o-(4-hydroxybenzoyl)benzoic acid, oxalic acid,
p-chlorobenzenesulfonic acid, phenyl-substituted alkanoic acids,
propionic acid, p-toluenesulfonic acid, pyruvic acid, salicylic
acid, stearic acid, succinic acid, tartaric acid,
tertiarybutylacetic acid, trimethylacetic acid, and the like.
Pharmaceutically acceptable salts also include base addition salts
which may be formed when acidic protons present are capable of
reacting with inorganic or organic bases. Acceptable inorganic
bases include sodium hydroxide, sodium carbonate, potassium
hydroxide, aluminum hydroxide and calcium hydroxide. Acceptable
organic bases include ethanolamine, diethanolamine,
triethanolamine, tromethamine, N-methylglucamine and the like. It
should be recognized that the particular anion or cation forming a
part of any salt of this invention is not critical, so long as the
salt, as a whole, is pharmacologically acceptable. Additional
examples of pharmaceutically acceptable salts and their methods of
preparation and use are presented in Handbook of Pharmaceutical
Salts: Properties, and Use (2002).
As used herein, "predominantly one enantiomer" means that a
compound contains at least about 85% of one enantiomer, or more
preferably at least about 90% of one enantiomer, or even more
preferably at least about 95% of one enantiomer, or most preferably
at least about 99% of one enantiomer. Similarly, the phrase
"substantially free from other optical isomers" means that the
composition contains at most about 15% of another enantiomer or
diastereomer, more preferably at most about 10% of another
enantiomer or diastereomer, even more preferably at most about 5%
of another enantiomer or diastereomer, and most preferably at most
about 1% of another enantiomer or diastereomer.
"Prevention" or "preventing" includes: (1) inhibiting the onset of
a disease in a subject or patient which may be at risk and/or
predisposed to the disease but does not yet experience or display
any or all of the pathology or symptomatology of the disease,
and/or (2) slowing the onset of the pathology or symptomatology of
a disease in a subject or patient which may be at risk and/or
predisposed to the disease but does not yet experience or display
any or all of the pathology or symptomatology of the disease.
"Prodrug" means a compound that is convertible in vivo
metabolically into an inhibitor according to the present
disclosure. The prodrug itself may or may not also have activity
with respect to a given target protein. For example, a compound
comprising a hydroxy group may be administered as an ester that is
converted by hydrolysis in vivo to the hydroxy compound. Suitable
esters that may be converted in vivo into hydroxy compounds include
acetates, citrates, lactates, phosphates, tartrates, malonates,
oxalates, salicylates, propionates, succinates, fumarates,
maleates, methylene-bis-.beta.-hydroxynaphthoate, gentisates,
isethionates, di-p-toluoyltartrates, methanesulfonates,
ethanesulfonates, benzenesulfonates, p-toluenesulfonates,
cyclohexylsulfamates, quinates, esters of amino acids, and the
like. Similarly, a compound comprising an amine group may be
administered as an amide that is converted by hydrolysis in vivo to
the amine compound.
The term "saturated" when referring to an atom means that the atom
is connected to other atoms only by means of single bonds.
A "stereoisomer" or "optical isomer" is an isomer of a given
compound in which the same atoms are bonded to the same other
atoms, but where the configuration of those atoms in three
dimensions differs. "Enantiomers" are stereoisomers of a given
compound that are mirror images of each other, like left and right
hands. "Diastereomers" are stereoisomers of a given compound that
are not enantiomers.
The invention contemplates that for any stereocenter or axis of
chirality for which stereochemistry has not been defined, that
stereocenter or axis of chirality can be present in its R form, S
form, or as a mixture of the R and S forms, including racemic and
non-racemic mixtures.
"Substituent convertible to hydrogen in vivo" means any group that
is convertible to a hydrogen atom by enzymological or chemical
means including, but not limited to, hydrolysis and hydrogenolysis.
Examples include acyl groups, groups having an oxycarbonyl group,
amino acid residues, peptide residues, o-nitrophenylsulfenyl,
trimethylsilyl, tetrahydro-pyranyl, diphenylphosphinyl, hydroxy or
alkoxy substituents on imino groups, and the like. Examples of acyl
groups include formyl, acetyl, trifluoroacetyl, and the like.
Examples of groups having an oxycarbonyl group include
ethoxycarbonyl, tert-butoxycarbonyl (--C(O)OC(CH.sub.3).sub.3),
benzyloxycarbonyl, p-methoxy-benzyloxycarbonyl, vinyloxycarbonyl,
.beta.-(p-toluenesulfonyl)ethoxycarbonyl, and the like. Suitable
amino acid residues include, but are not limited to, residues of
Gly (glycine), Ala (alanine), Arg (arginine), Asn (asparagine), Asp
(aspartic acid), Cys (cysteine), Glu (glutamic acid), His
(histidine), Ile (isoleucine), Leu (leucine), Lys (lysine), Met
(methionine), Phe (phenylalanine), Pro (proline), Ser (serine), Thr
(threonine), Trp (tryptophan), Tyr (tyrosine), Val (valine), Nva
(norvaline), Hse (homoserine), 4-Hyp (4-hydroxyproline), 5-Hyl
(5-hydroxylysine), Orn (ornithine) and .beta.-Ala. Examples of
suitable amino acid residues also include amino acid residues that
are protected with a protecting group. Examples of suitable
protecting groups include those typically employed in peptide
synthesis, including acyl groups (such as formyl and acetyl),
arylmethyloxycarbonyl groups (such as benzyloxycarbonyl and
p-nitrobenzyloxycarbonyl), tert-butoxycarbonyl groups
(--C(O)OC(CH.sub.3).sub.3), and the like. Suitable peptide residues
include peptide residues comprising two to five, and optionally
amino acid residues. The residues of these amino acids or peptides
can be present in stereochemical configurations of the D-form, the
L-form or mixtures thereof. In addition, the amino acid or peptide
residue may have an asymmetric carbon atom. Examples of suitable
amino acid residues having an asymmetric carbon atom include
residues of Ala, Leu, Phe, Trp, Nva, Val, Met, Ser, Lys, Thr and
Tyr. Peptide residues having an asymmetric carbon atom include
peptide residues having one or more constituent amino acid residues
having an asymmetric carbon atom. Examples of suitable amino acid
protecting groups include those typically employed in peptide
synthesis, including acyl groups (such as formyl and acetyl),
arylmethyloxycarbonyl groups (such as benzyloxycarbonyl and
p-nitrobenzyloxycarbonyl), tert-butoxycarbonyl groups
(--C(O)OC(CH.sub.3).sub.3), and the like. Other examples of
substituents "convertible to hydrogen in vivo" include reductively
eliminable hydrogenolyzable groups. Examples of suitable
reductively eliminable hydrogenolyzable groups include, but are not
limited to, arylsulfonyl groups (such as o-toluenesulfonyl); methyl
groups substituted with phenyl or benzyloxy (such as benzyl, trityl
and benzyloxymethyl); arylmethoxycarbonyl groups (such as
benzyloxycarbonyl and o-methoxy-benzyloxycarbonyl); and
haloethoxycarbonyl groups (such as
.beta.,.beta.,.beta.-trichloroethoxycarbonyl and
.beta.-iodoethoxycarbonyl).
"Therapeutically effective amount" or "pharmaceutically effective
amount" means that amount which, when administered to a subject or
patient for treating a disease, is sufficient to effect such
treatment for the disease.
"Treatment" or "treating" includes (1) inhibiting a disease in a
subject or patient experiencing or displaying the pathology or
symptomatology of the disease (e.g., arresting further development
of the pathology and/or symptomatology), (2) ameliorating a disease
in a subject or patient that is experiencing or displaying the
pathology or symptomatology of the disease (e.g., reversing the
pathology and/or symptomatology), and/or (3) effecting any
measurable decrease in a disease in a subject or patient that is
experiencing or displaying the pathology or symptomatology of the
disease.
As used herein, the term "water soluble" means that the compound
dissolves in water at least to the extent of 0.010 mole/liter or is
classified as soluble according to literature precedence.
Other abbreviations used herein are as follows: DMSO, dimethyl
sulfoxide; NO, nitric oxide; iNOS, inducible nitric oxide synthase;
COX-2, cyclooxygenase-2; NGF, nerve growth factor; IBMX,
isobutylmethylxanthine; FBS, fetal bovine serum; GPDH, glycerol
3-phosphate dehydrogenase; RXR, retinoid X receptor; TGF-.beta.,
transforming growth factor-.beta.; IFN.gamma. or IFN-.gamma.,
interferon-.gamma.; LPS, bacterial endotoxic lipopolysaccharide;
TNF.alpha. or TNF-.alpha., tumor necrosis factor-.alpha.;
IL-1.beta., interleukin-1.beta.; GAPDH, glyceraldehyde-3-phosphate
dehydrogenase; MTT,
3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide; TCA,
trichloroacetic acid; HO-1, inducible heme oxygenase.
The above definitions supersede any conflicting definition in any
of the reference that is incorporated by reference herein. The fact
that certain terms are defined, however, should not be considered
as indicative that any term that is undefined is indefinite.
Rather, all terms used are believed to describe the invention in
terms such that one of ordinary skill can appreciate the scope and
practice the present disclosure.
II. SYNTHETIC METHODS
Compounds of the present disclosure may be made using the methods
outlined in the Examples section (Example 2 and 3). These methods
can be further modified and optimized using the principles and
techniques of organic chemistry as applied by a person skilled in
the art. Such principles and techniques are taught, for example, in
March's Advanced Organic Chemistry: Reactions, Mechanisms, and
Structure (2007), which is incorporated by reference herein.
III. BIOLOGICAL ACTIVITY OF OLEANOLIC ACID DERIVATIVES
Biological activity results, both in vivo and in vitro are provided
throughout the present disclosure. These include: inhibition of NO
Production, suppression of COX-2 induction, inhibition of IL-6
induced STAT3 phosphorylation, suppression of IL-6 induced STAT3
phosphorylation, inhibition of TNF.alpha.-induced I.kappa.B.alpha.
degradation, inhibition of NF.kappa.B activation, induction of
HO-1, Nrf2 induction of HO-1, TrxR1 and .gamma.-GCS, induction of
TrxR1, induction of .gamma.-GCS, induction of ferritin heavy chain,
induction of TrxR1, induction of .gamma.-GCS, induction of ferritin
heavy chain, and various in vivo toxicity studies. See figures and
figure descriptions. Suppression of NO production and induction of
Nrf2 induction results can be respectively summarized as shown
Tables 1a and 1b, below. Further results, including toxicity
studies, are provided in the Examples section.
TABLE-US-00001 TABLE 1a Suppression of IFN.gamma.-Induced NO
Production. RAW264.7 (20 ng/ml IFN.gamma.) iNOS suppr. Compound
ID(s) MW NO IC.sub.50 WST-1 IC.sub.50 WB 63101/402-02/dh402 507.70
~12 nM 200 nM >90% 63102/404-02/dh404 574.72 ~45 nM >200 nM
63250/402-46 509.70 >200 nM >200 nM 63197/402-48 512.72
>200 nM >200 nM 63195/402-49 509.72 >200 nM >200 nM
63196/402-51/dh401 493.68 ~75 nM >200 nM 63252/402-57 474.68 ~5
nM >200 nM 63205/402-59 492.69 ~25 nM >200 nM 63206/402-64
477.68 ~10 nM >200 nM 63207/402-66 509.72 ~50 nM >200 nM
63219/402-78 509.72 ~150 nM >200 nM 63229 517.71 >200 nM
>200 nM 63230 531.74 ~50 nM >200 nM 63227 576.73 >200 nM
>200 nM 63219 509.72 ~150 nM >200 nM 63223 576.73 >200 nM
>200 nM 63237 572.70 ~80 nM >200 nM 63268 576.75 >200 nM
>200 nM 63274 547.81 ~70 nM >200 nM 63289 525.73 >200 nM
>200 nM 63295 522.73 ~20 nM >200 nM 63296 522.73 ~200 nM
>200 nM 63308 491.70 ~25 nM >200 nM 63323 583.80 ~40 nM See
FIG. 33 63325 520.75 ~50 nM See FIG. 32 63326 552.79 ~50 nM See
FIG. 34
TABLE-US-00002 TABLE 1b Induction of HO-1, TrxR1 and .gamma.-GCS in
Human Melanoma Cells. Nrf2 target gene induction in MDA-MB-435
cells Compound 400 nM* 250 nM** Code HO-1 TrxR1 .gamma.-GCS HO-1
NQO1 .gamma.-GCS 63101 4 47 53 3 2 5 (dh402) 63102 3 2 4.5 (dh404)
63196 1 48 26 (dh401) 63252 5 3 63205 1.7 1.7 3.5 63206 2.5 1.8 4.5
63207 1.2 1.6 3.1 63237 3 2.3 6.4 Blank entry: Not determined.
*Data expressed as a percent of induction observed for 402 (see
below for structure). **Data expressed as fold induction above DMSO
control.
In certain embodiments, the compounds of the present disclosure are
capable of crossing the blood brain barrier and achieving
therapeutically effective concentrations in the brain. They may
therefore be used to treat neurodegenerative diseases, brain cancer
and other inflammatory conditions affecting the central nervous
system. For example, 404-02 has been shown to cross the blood-brain
barrier and achieve high concentrations in the central nervous
system tissue following oral dosing. Like the other compounds of
the present disclosure, it promotes the resolution of innate and
adaptive immune-mediated inflammation by restoring redox
homeostasis in inflamed tissues. It is a potent inducer of the
antioxidant transcription factor Nrf2 and inhibitor of the
pro-oxidant/pro-inflammatory transcription factors NF-.kappa.B and
the STATs. These biological pathways are implicated in a wide
variety of diseases, including autoimmune conditions and several
neurodegenerative diseases.
IV. IMPROVED RODENT TOXICOLOGY
In certain embodiments, the invention provides compounds possessing
low toxicity in rodents. In some cases, toxicity in rodents has
been observed in preclinical studies with some analogues containing
a carbon-carbon double bond in the C-ring, including 402 and 401.
Compounds having a saturated C-ring, in contrast, have consistently
shown low toxicity in rodents. Predictably low rodent toxicity
provides an advantage since high rodent toxicity can be a
significant complication in conducting preclinical studies required
for development and registration of therapeutic compounds for use
in humans or non-human animals. Illustrations of this effect are
provided below.
##STR00038##
For example, an initial study (Example 6) was performed in Sprague
Dawley rats using both 402 and 402-02 and showed that 402-02 was
less toxic. In a further study (Example 7), six compounds (401,
402, 404, 401-2, 402-2, and 404-2) were assessed for toxicity in
mice in a 14-day study. At higher doses (above 10 mg/kg/day) both
401 and 402 caused at least 50% mortality, while 404 was non-toxic.
In contrast, no mortality was observed in the 402-2 and 404-2
groups and only the highest dose of 401-02 caused any lethality
(Table 5). Body weight measurements (FIGS. 29-31) were consistent
with the mortality observations. Notably, the two highest doses of
401 and 402 were lethal within 4 days, in contrast to the effects
of 401-2 and 402-2.
V. DISEASES ASSOCIATED WITH INFLAMMATION AND/OR OXIDATIVE
STRESS
Inflammation is a biological process that provides resistance to
infectious or parasitic organisms and the repair of damaged tissue.
Inflammation is commonly characterized by localized vasodilation,
redness, swelling, and pain, the recruitment of leukocytes to the
site of infection or injury, production of inflammatory cytokines
such as TNF-.alpha. and IL-1, and production of reactive oxygen or
nitrogen species such as hydrogen peroxide, superoxide and
peroxynitrite. In later stages of inflammation, tissue remodeling,
angiogenesis, and scar formation (fibrosis) may occur as part of
the wound healing process. Under normal circumstances, the
inflammatory response is regulated and temporary and is resolved in
an orchestrated fashion once the infection or injury has been dealt
with adequately. However, acute inflammation can become excessive
and life-threatening if regulatory mechanisms fail. Alternatively,
inflammation can become chronic and cause cumulative tissue damage
or systemic complications.
Many serious and intractable human diseases involve dysregulation
of inflammatory processes, including diseases such as cancer,
atherosclerosis, and diabetes, which were not traditionally viewed
as inflammatory conditions. In the case of cancer, the inflammatory
processes are associated with tumor formation, progression,
metastasis, and resistance to therapy. Atherosclerosis, long viewed
as a disorder of lipid metabolism, is now understood to be
primarily an inflammatory condition, with activated macrophages
playing an important role in the formation and eventual rupture of
atherosclerotic plaques. Activation of inflammatory signaling
pathways has also been shown to play a role in the development of
insulin resistance, as well as in the peripheral tissue damage
associated with diabetic hyperglycemia. Excessive production of
reactive oxygen species and reactive nitrogen species such as
superoxide, hydrogen peroxide, nitric oxide, and peroxynitrite is a
hallmark of inflammatory conditions. Evidence of dysregulated
peroxynitrite production has been reported in a wide variety of
diseases (Szabo et al., 2007; Schulz et al., 2008; Forstermann,
2006; Pall, 2007).
Autoimmune diseases such as rheumatoid arthritis, lupus, psoriasis,
and multiple sclerosis involve inappropriate and chronic activation
of inflammatory processes in affected tissues, arising from
dysfunction of self vs. non-self recognition and response
mechanisms in the immune system. In neurodegenerative diseases such
as Alzheimer's and Parkinson's diseases, neural damage is
correlated with activation of microglia and elevated levels of
pro-inflammatory proteins such as inducible nitric oxide synthase
(iNOS). Chronic organ failure such as renal failure, heart failure,
and chronic obstructive pulmonary disease is closely associated
with the presence of chronic oxidative stress and inflammation,
leading to the development of fibrosis and eventual loss of organ
function.
Many other disorders involve oxidative stress and inflammation in
affected tissues, including inflammatory bowel disease;
inflammatory skin diseases; mucositis related to radiation therapy
and chemotherapy; eye diseases such as uveitis, glaucoma, macular
degeneration, and various forms of retinopathy; transplant failure
and rejection; ischemia-reperfusion injury; chronic pain;
degenerative conditions of the bones and joints including
osteoarthritis and osteoporosis; asthma and cystic fibrosis;
seizure disorders; and neuropsychiatric conditions including
schizophrenia, depression, bipolar disorder, post-traumatic stress
disorder, attention deficit disorders, autism-spectrum disorders,
and eating disorders such as anorexia nervosa. Dysregulation of
inflammatory signaling pathways is believed to be a major factor in
the pathology of muscle wasting diseases including muscular
dystrophy and various forms of cachexia.
A variety of life-threatening acute disorders also involve
dysregulated inflammatory signaling, including acute organ failure
involving the pancreas, kidneys, liver, or lungs, myocardial
infarction or acute coronary syndrome, stroke, septic shock,
trauma, severe burns, and anaphylaxis.
Many complications of infectious diseases also involve
dysregulation of inflammatory responses. Although an inflammatory
response can kill invading pathogens, an excessive inflammatory
response can also be quite destructive and in some cases can be a
primary source of damage in infected tissues. Furthermore, an
excessive inflammatory response can also lead to systemic
complications due to overproduction of inflammatory cytokines such
as TNF-.alpha. and IL-1. This is believed to be a factor in
mortality arising from severe influenza, severe acute respiratory
syndrome, and sepsis.
The aberrant or excessive expression of either iNOS or
cyclooxygenase-2 (COX-2) has been implicated in the pathogenesis of
many disease processes. For example, it is clear that NO is a
potent mutagen (Tamir and Tannebaum, 1996), and that nitric oxide
can also activate COX-2 (Salvemini et al., 1994). Furthermore,
there is a marked increase in iNOS in rat colon tumors induced by
the carcinogen, azoxymethane (Takahashi et al., 1997). A series of
synthetic triterpenoid analogs of oleanolic acid have been shown to
be powerful inhibitors of cellular inflammatory processes, such as
the induction by IFN-.gamma. of inducible nitric oxide synthase
(iNOS) and of COX-2 in mouse macrophages. See Honda et al. (2000a);
Honda et al. (2000b), and Honda et al. (2002), which are all
incorporated herein by reference.
In one aspect, compounds of the invention are characterized by
their ability to inhibit the production of nitric oxide in
macrophage-derived RAW 264.7 cells induced by exposure to
.gamma.-interferon. They are further characterized by their ability
to induce the expression of antioxidant proteins such as NQO1 and
reduce the expression of pro-inflammatory proteins such as COX-2
and inducible nitric oxide synthase (iNOS). These properties are
relevant to the treatment of a wide array of diseases involving
oxidative stress and dysregulation of inflammatory processes
including cancer, mucositis resulting from radiation therapy or
chemotherapy, autoimmune diseases, cardiovascular diseases
including atherosclerosis, ischemia-reperfusion injury, acute and
chronic organ failure including renal failure and heart failure,
respiratory diseases, diabetes and complications of diabetes,
severe allergies, transplant rejection, graft-versus-host disease,
neurodegenerative diseases, diseases of the eye and retina, acute
and chronic pain, degenerative bone diseases including
osteoarthritis and osteoporosis, inflammatory bowel diseases,
dermatitis and other skin diseases, sepsis, burns, seizure
disorders, and neuropsychiatric disorders.
Without being bound by theory, the activation of the
antioxidant/anti-inflammatory Keap1/Nrf2/ARE pathway is believed to
be implicated in both the anti-inflammatory and anti-carcinogenic
properties of the present oleanolic acid derivatives.
In another aspect, compounds of the invention may be used for
treating a subject having a condition caused by elevated levels of
oxidative stress in one or more tissues. Oxidative stress results
from abnormally high or prolonged levels of reactive oxygen species
such as superoxide, hydrogen peroxide, nitric oxide, and
peroxynitrite (formed by the reaction of nitric oxide and
superoxide). The oxidative stress may be accompanied by either
acute or chronic inflammation. The oxidative stress may be caused
by mitochondrial dysfunction, by activation of immune cells such as
macrophages and neutrophils, by acute exposure to an external agent
such as ionizing radiation or a cytotoxic chemotherapy agent (e.g.,
doxorubicin), by trauma or other acute tissue injury, by
ischemia/reperfusion, by poor circulation or anemia, by localized
or systemic hypoxia or hyperoxia, by elevated levels of
inflammatory cytokines and other inflammation-related proteins,
and/or by other abnormal physiological states such as hyperglycemia
or hypoglycemia.
In animal models of many such conditions, stimulating expression of
inducible heme oxygenase (HO-1), a target gene of the Nrf2 pathway,
has been shown to have a significant therapeutic effect including
models of myocardial infarction, renal failure, transplant failure
and rejection, stroke, cardiovascular disease, and autoimmune
disease (e.g., Sacerdoti et al., 2005; Abraham & Kappas, 2005;
Bach, 2006; Araujo et al., 2003; Liu et al., 2006; Ishikawa et al.,
2001; Kruger et al., 2006; Satoh et al., 2006; Zhou et al., 2005;
Morse and Choi, 2005; Morse and Choi, 2002). This enzyme breaks
free heme down into iron, carbon monoxide (CO), and biliverdin
(which is subsequently converted to the potent antioxidant
molecule, bilirubin).
In another aspect, compounds of this invention may be used in
preventing or treating tissue damage or organ failure, acute and
chronic, resulting from oxidative stress exacerbated by
inflammation. Examples of diseases that fall in this category
include: heart failure, liver failure, transplant failure and
rejection, renal failure, pancreatitis, fibrotic lung diseases
(cystic fibrosis and COPD, among others), diabetes (including
complications), atherosclerosis, ischemia-reperfusion injury,
glaucoma, stroke, autoimmune disease, autism, macular degeneration,
and muscular dystrophy. For example, in the case of autism, studies
suggest that increased oxidative stress in the central nervous
system may contribute to the development of the disease (Chauhan
and Chauhan, 2006).
Evidence also links oxidative stress and inflammation to the
development and pathology of many other disorders of the central
nervous system, including psychiatric disorders such as psychosis,
major depression, and bipolar disorder; seizure disorders such as
epilepsy; pain and sensory syndromes such as migraine, neuropathic
pain or tinnitus; and behavioral syndromes such as the attention
deficit disorders. See, e.g., Dickerson et al., 2007; Hanson et
al., 2005; Kendall-Tackett, 2007; Lencz et al., 2007; Dudhgaonkar
et al., 2006; Lee et al., 2007; Morris et al., 2002; Ruster et al.,
2005; McIver et al., 2005; Sarchielli et al., 2006; Kawakami et
al., 2006; Ross et al., 2003, which are all incorporated by
reference herein. For example, elevated levels of inflammatory
cytokines, including TNF, interferon-.gamma., and IL-6, are
associated with major mental illness (Dickerson et al., 2007).
Microglial activation has also been linked to major mental illness.
Therefore, downregulating inflammatory cytokines and inhibiting
excessive activation of microglia could be beneficial in patients
with schizophrenia, major depression, bipolar disorder,
autism-spectrum disorders, and other neuropsychiatric
disorders.
Accordingly, in pathologies involving oxidative stress alone or
oxidative stress exacerbated by inflammation, treatment may
comprise administering to a subject a therapeutically effective
amount of a compound of this invention, such as those described
above or throughout this specification. Treatment may be
administered preventively, in advance of a predictable state of
oxidative stress (e.g., organ transplantation or the administration
of radiation therapy to a cancer patient), or it may be
administered therapeutically in settings involving established
oxidative stress and inflammation.
The compounds of the invention may be generally applied to the
treatment of inflammatory conditions, such as sepsis, dermatitis,
autoimmune disease and osteoarthritis. In one aspect, the compounds
of this invention may be used to treat inflammatory pain and/or
neuropathic pain, for example, by inducing Nrf2 and/or inhibiting
NF-.kappa.B.
In one aspect, the compounds of the invention may be used to
function as antioxidant inflammation modulators (AIMs) having
potent anti-inflammatory properties that mimic the biological
activity of cyclopentenone prostaglandins (cyPGs). In one
embodiment, the compounds of the invention may be used to control
the production of pro-inflammatory cytokines by selectively
targeting regulatory cysteine residues (RCRs) on proteins that
regulate the transcriptional activity of redox-sensitive
transcription factors. Activation of RCRs by cyPGs or AIMs has been
shown to initiate a pro-resolution program in which the activity of
the antioxidant and cytoprotective transcription factor Nrf2 is
potently induced, and the activities of the pro-oxidant and
pro-inflammatory transcription factors NF-.kappa.B and the STATs
are suppressed. This increases the production of antioxidant and
reductive molecules (e.g., NQO1, HO-1, SOD1, and/or .gamma.-GCS)
and/or decreases oxidative stress and the production of pro-oxidant
and pro-inflammatory molecules (e.g., iNOS, COX-2, and/or
TNF-.alpha.).
In some embodiments, the compounds of the invention may be used in
the treatment and prevention of diseases such as cancer,
inflammation, Alzheimer's disease, Parkinson's disease, multiple
sclerosis, autism, amyotrophic lateral sclerosis, autoimmune
diseases such as rheumatoid arthritis, lupus, and MS, inflammatory
bowel disease, all other diseases whose pathogenesis is believed to
involve excessive production of either nitric oxide or
prostaglandins, and pathologies involving oxidative stress alone or
oxidative stress exacerbated by inflammation.
Another aspect of inflammation is the production of inflammatory
prostaglandins such as prostaglandin E. These molecules promote
vasodilation, plasma extravasation, localized pain, elevated
temperature, and other symptoms of inflammation. The inducible form
of the enzyme COX-2 is associated with their production, and high
levels of COX-2 are found in inflamed tissues. Consequently,
inhibition of COX-2 may relieve many symptoms of inflammation and a
number of important anti-inflammatory drugs (e.g., ibuprofen and
celecoxib) act by inhibiting COX-2 activity. Recent research,
however, has demonstrated that a class of cyclopentenone
prostaglandins (cyPGs) (e.g., 15-deoxy prostaglandin J2, a.k.a.
PGJ2) plays a role in stimulating the orchestrated resolution of
inflammation (e.g., Rajakariar et al., 2007). COX-2 is also
associated with the production of cyclopentenone prostaglandins.
Consequently, inhibition of COX-2 may interfere with the full
resolution of inflammation, potentially promoting the persistence
of activated immune cells in tissues and leading to chronic,
"smoldering" inflammation. This effect may be responsible for the
increased incidence of cardiovascular disease in patients using
selective COX-2 inhibitors for long periods of time.
In one aspect, the compounds of the invention may be used to
control the production of pro-inflammatory cytokines within the
cell by selectively activating regulatory cysteine residues (RCRs)
on proteins that regulate the activity of redox-sensitive
transcription factors. Activation of RCRs by cyPGs has been shown
to initiate a pro-resolution program in which the activity of the
antioxidant and cytoprotective transcription factor Nrf2 is
potently induced and the activities of the pro-oxidant and
pro-inflammatory transcription factors NF-.kappa.B and the STATs
are suppressed. In some embodiments, this increases the production
of antioxidant and reductive molecules (NQO1, HO-1, SOD1,
.gamma.-GCS) and decreases oxidative stress and the production of
pro-oxidant and pro-inflammatory molecules (iNOS, COX-2,
TNF-.alpha.). In some embodiments, the compounds of this invention
may cause the cells that host the inflammatory event to revert to a
non-inflammatory state by promoting the resolution of inflammation
and limiting excessive tissue damage to the host.
A. Cancer
Further, the compounds of the present disclosure may be used to
induce apoptosis in tumor cells, to induce cell differentiation, to
inhibit cancer cell proliferation, to inhibit an inflammatory
response, and/or to function in a chemopreventative capacity. For
example, the invention provides new compounds that have one or more
of the following properties: (1) an ability to induce apoptosis and
differentiate both malignant and non-malignant cells, (2) an
activity at sub-micromolar or nanomolar levels as an inhibitor of
proliferation of many malignant or premalignant cells, (3) an
ability to suppress the de novo synthesis of the inflammatory
enzyme inducible nitric oxide synthase (iNOS), (4) an ability to
inhibit NF-.kappa.B activation, and (5) an ability to induce the
expression of heme oxygenase-1 (HO-1).
The levels of iNOS and COX-2 are elevated in certain cancers and
have been implicated in carcinogenesis and COX-2 inhibitors have
been shown to reduce the incidence of primary colonic adenomas in
humans (Rostom et al., 2007; Brown and DuBois, 2005; Crowel et al.,
2003). iNOS is expressed in myeloid-derived suppressor cells
(MDSCs) (Angulo et al., 2000) and COX-2 activity in cancer cells
has been shown to result in the production of prostaglandin E.sub.2
(PGE.sub.2), which has been shown to induce the expression of
arginase in MDSCs (Sinha et al., 2007). Arginase and iNOS are
enzymes that utilize L-arginine as a substrate and produce
L-ornithine and urea, and L-citrulline and NO, respectively. The
depletion of arginine from the tumor microenvironment by MDSCs,
combined with the production of NO and peroxynitrite has been shown
to inhibit proliferation and induce apoptosis of T cells (Bronte et
al., 2003) Inhibition of COX-2 and iNOS has been shown to reduce
the accumulation of MDSCs, restore cytotoxic activity of
tumor-associated T cells, and delay tumor growth (Sinha et al.,
2007; Mazzoni et al., 2002; Zhou et al., 2007).
Inhibition of the NF-.kappa.B and JAK/STAT signaling pathways has
been implicated as a strategy to inhibit proliferation of cancer
epithelial cells and induce their apoptosis. Activation of STAT3
and NF-.kappa.B has been shown to result in suppression of
apoptosis in cancer cells, and promotion of proliferation,
invasion, and metastasis. Many of the target genes involved in
these processes have been shown to be transcriptionally regulated
by both NF-.kappa.B and STAT3 (Yu et al., 2007).
In addition to their direct roles in cancer epithelial cells,
NF-.kappa.B and STAT3 also have important roles in other cells
found within the tumor microenvironment. Experiments in animal
models have demonstrated that NF-.kappa.B is required in both
cancer cells and hematopoeitic cells to propagate the effects of
inflammation on cancer initiation and progression (Greten et al.,
2004). NF-.kappa.B inhibition in cancer and myeloid cells reduces
the number and size, respectively, of the resultant tumors.
Activation of STAT3 in cancer cells results in the production of
several cytokines (IL-6, IL-10) which suppress the maturation of
tumor-associated dendritic cells (DC). Furthermore, STAT3 is
activated by these cytokines in the dendritic cells themselves
Inhibition of STAT3 in mouse models of cancer restores DC
maturation, promotes antitumor immunity, and inhibits tumor growth
(Kortylewski et al., 2005).
B. Treatment of Multiple Sclerosis and Other Neurodegenerative
Conditions
The compounds and methods of this invention may be used for
treating patients for multiple sclerosis (MS). MS is known to be an
inflammatory condition of the central nervous system (Williams et
al., 1994; Merrill and Benvenist, 1996; Genain and Nauser, 1997).
Based on several investigations, there is evidence suggesting that
inflammatory, oxidative, and/or immune mechanisms are involved in
the pathogenesis of Alzheimer's disease (AD), Parkinson's disease
(PD), amyotrophic lateral sclerosis (ALS), and MS (Bagasra et al.,
1995; McGeer and McGeer, 1995; Simonian and Coyle, 1996;
Kaltschmidt et al., 1997). Both reactive astrocytes and activated
microglia have been implicated in causation of neurodegenerative
disease (NDD) and neuroinflammatory disease (NID); there has been a
particular emphasis on microglia as cells that synthesize both NO
and prostaglandins as products of the respective enzymes, iNOS and
COX-2. De novo formation of these enzymes may be driven by
inflammatory cytokines such as interferon-.gamma. or interleukin-1.
In turn, excessive production of NO may lead to inflammatory
cascades and/or oxidative damage in cells and tissues of many
organs, including neurons and oligodendrocytes of the nervous
system, with consequent manifestations in AD and MS, and possible
PD and ALS (Coyle and Puttfarcken, 1993; Beal, 1996; Merrill and
Benvenist, 1996; Simonian and Coyle, 1996; Vodovotz et al., 1996).
Epidemiologic data indicate that chronic use of NSAID's which block
synthesis of prostaglandins from arachidonate, markedly lower the
risk for development of AD (McGeer et al., 1996; Stewart et al.,
1997). Thus, agents that block formation of NO and prostaglandins,
may be used in approaches to prevention and treatment of NDD.
Successful therapeutic candidates for treating such a disease
typically require an ability to penetrate the blood-brain barrier.
See, for example, U.S. Patent Publication 2009/0060873, which is
incorporated by reference herein in its entirety. See also, for
example, the results presented for compound 404-02 in Examples 4
and 5, below.
C. Neuroinflammation
The compounds and methods of this invention may be used for
treating patients with neuroinflammation. Neuroinflammation
encapsulates the idea that microglial and astrocytic responses and
actions in the central nervous system have a fundamentally
inflammation-like character, and that these responses are central
to the pathogenesis and progression of a wide variety of
neurological disorders. This idea originated in the field of
Alzheimer's disease (Griffin et al., 1989; Rogers et al., 1988),
where it has revolutionized our understanding of this disease
(Akiyama et al., 2000). These ideas have been extended to other
neurodegenerative diseases (Eikelenboom et al., 2002; Ishizawa and
Dickson, 2001), to ischemic/toxic diseases (Gehrmann et al., 1995;
Touzani et al., 1999), to tumor biology (Graeber et al., 2002) and
even to normal brain development.
Neuroinflammation incorporates a wide spectrum of complex cellular
responses that include activation of microglia and astrocytes and
induction of cytokines, chemokines, complement proteins, acute
phase proteins, oxidative injury, and related molecular processes.
These events may have detrimental effects on neuronal function,
leading to neuronal injury, further glial activation, and
ultimately neurodegeneration.
D. Treatment of Renal Failure
The compounds and methods of this invention may be used for
treating patients with renal failure. See U.S. patent application
Ser. No. 12/352,473, which is incorporated by reference herein in
its entirety. Another aspect of the present disclosure concerns new
methods and compounds for the treatment and prevention of renal
disease. Renal failure, resulting in inadequate clearance of
metabolic waste products from the blood and abnormal concentrations
of electrolytes in the blood, is a significant medical problem
throughout the world, especially in developed countries. Diabetes
and hypertension are among the most important causes of chronic
renal failure, also known as chronic kidney disease (CKD), but it
is also associated with other conditions such as lupus. Acute renal
failure may arise from exposure to certain drugs (e.g.,
acetaminophen) or toxic chemicals, or from ischemia-reperfusion
injury associated with shock or surgical procedures such as
transplantation, and may result in chronic renal failure. In many
patients, renal failure advances to a stage in which the patient
requires regular dialysis or kidney transplantation to continue
living. Both of these procedures are highly invasive and associated
with significant side effects and quality of life issues. Although
there are effective treatments for some complications of renal
failure, such as hyperparathyroidism and hyperphosphatemia, no
available treatment has been shown to halt or reverse the
underlying progression of renal failure. Thus, agents that can
improve compromised renal function would represent a significant
advance in the treatment of renal failure.
Inflammation contributes significantly to the pathology of CKD.
There is also a strong mechanistic link between oxidative stress
and renal dysfunction. The NF-.kappa.B signaling pathway plays an
important role in the progression of CKD as NF-.kappa.B regulates
the transcription of MCP-1, a chemokine that is responsible for the
recruitment of monocytes/macrophages resulting in an inflammatory
response that ultimately injures the kidney (Wardle, 2001). The
Keap1/Nrf2/ARE pathway controls the transcription of several genes
encoding antioxidant enzymes, including heme oxygenase-1 (HO-1).
Ablation of the Nrf2 gene in female mice results in the development
of lupus-like glomerular nephritis (Yoh et al., 2001). Furthermore,
several studies have demonstrated that HO-1 expression is induced
in response to renal damage and inflammation and that this enzyme
and its products--bilirubin and carbon monoxide--play a protective
role in the kidney (Nath et al., 2006).
The glomerulus and the surrounding Bowman's capsule constitute the
basic functional unit of the kidney. Glomerular filtration rate
(GFR) is the standard measure of renal function. Creatinine
clearance is commonly used to measure GFR. However, the level of
serum creatinine is commonly used as a surrogate measure of
creatinine clearance. For instance, excessive levels of serum
creatinine are generally accepted to indicate inadequate renal
function and reductions in serum creatinine over time are accepted
as an indication of improved renal function. Normal levels of
creatinine in the blood are approximately 0.6 to 1.2 milligrams
(mg) per deciliter (dl) in adult males and 0.5 to 1.1 milligrams
per deciliter in adult females.
Acute kidney injury (AKI) can occur following ischemia-reperfusion,
treatment with certain pharmacological agents such as cisplatin and
rapamycin, and intravenous injection of radiocontrast media used in
medical imaging. As in CKD, inflammation and oxidative stress
contribute to the pathology of AKI. The molecular mechanisms
underlying radiocontrast-induced nephropathy (RCN) are not well
understood; however, it is likely that a combination of events
including prolonged vasoconstriction, impaired kidney
autoregulation, and direct toxicity of the contrast media all
contribute to renal failure (Tumlin et al., 2006). Vasoconstriction
results in decreased renal blood flow and causes
ischemia-reperfusion and the production of reactive oxygen species.
HO-1 is strongly induced under these conditions and has been
demonstrated to prevent ischemia-reperfusion injury in several
different organs, including the kidney (Nath et al., 2006).
Specifically, induction of HO-1 has been shown to be protective in
a rat model of RCN (Goodman et al., 2007). Reperfusion also induces
an inflammatory response, in part though activation of NF-.kappa.B
signaling (Nichols, 2004). Targeting NF-.kappa.B has been proposed
as a therapeutic strategy to prevent organ damage (Zingarelli et
al., 2003).
E. Cardiovascular Disease
The compounds and methods of this invention may be used for
treating patients with cardiovascular disease. See U.S. patent
application Ser. No. 12/352,473, which is incorporated by reference
herein in its entirety. Cardiovascular (CV) disease is among the
most important causes of mortality worldwide, and is the leading
cause of death in many developed nations. The etiology of CV
disease is complex, but the majority of causes are related to
inadequate or completely disrupted supply of blood to a critical
organ or tissue. Frequently such a condition arises from the
rupture of one or more atherosclerotic plaques, which leads to the
formation of a thrombus that blocks blood flow in a critical
vessel. Such thrombosis is the principal cause of heart attacks, in
which one or more of the coronary arteries is blocked and blood
flow to the heart itself is disrupted. The resulting ischemia is
highly damaging to cardiac tissue, both from lack of oxygen during
the ischemic event and from excessive formation of free radicals
after blood flow is restored (a phenomenon known as
ischemia-reperfusion injury). Similar damage occurs in the brain
during a thrombotic stroke, when a cerebral artery or other major
vessel is blocked by thrombosis. Hemorrhagic strokes, in contrast,
involve rupture of a blood vessel and bleeding into the surrounding
brain tissue. This creates oxidative stress in the immediate area
of the hemorrhage, due to the presence of large amounts of free
heme and other reactive species, and ischemia in other parts of the
brain due to compromised blood flow. Subarachnoid hemorrhage, which
is frequently accompanied by cerebral vasospasm, also causes
ischemia/reperfusion injury in the brain.
Alternatively, atherosclerosis may be so extensive in critical
blood vessels that stenosis (narrowing of the arteries) develops
and blood flow to critical organs (including the heart) is
chronically insufficient. Such chronic ischemia can lead to
end-organ damage of many kinds, including the cardiac hypertrophy
associated with congestive heart failure.
Atherosclerosis, the underlying defect leading to many forms of
cardiovascular disease, occurs when a physical defect or injury to
the lining (endothelium) of an artery triggers an inflammatory
response involving the proliferation of vascular smooth muscle
cells and the infiltration of leukocytes into the affected area.
Ultimately, a complicated lesion known as an atherosclerotic plaque
may form, composed of the above-mentioned cells combined with
deposits of cholesterol-bearing lipoproteins and other materials
(e.g., Hansson et al., 2006).
Pharmaceutical treatments for cardiovascular disease include
preventive treatments, such as the use of drugs intended to lower
blood pressure or circulating levels of cholesterol and
lipoproteins, as well as treatments designed to reduce the adherent
tendencies of platelets and other blood cells (thereby reducing the
rate of plaque progression and the risk of thrombus formation).
More recently, drugs such as streptokinase and tissue plasminogen
activator have been introduced and are used to dissolve the
thrombus and restore blood flow. Surgical treatments include
coronary artery bypass grafting to create an alternative blood
supply, balloon angioplasty to compress plaque tissue and increase
the diameter of the arterial lumen, and carotid endarterectomy to
remove plaque tissue in the carotid artery. Such treatments,
especially balloon angioplasty, may be accompanied by the use of
stents, expandable mesh tubes designed to support the artery walls
in the affected area and keep the vessel open. Recently, the use of
drug-eluting stents has become common in order to prevent
post-surgical restenosis (renarrowing of the artery) in the
affected area. These devices are wire stents coated with a
biocompatible polymer matrix containing a drug that inhibits cell
proliferation (e.g., paclitaxel or rapamycin). The polymer allows a
slow, localized release of the drug in the affected area with
minimal exposure of non-target tissues. Despite the significant
benefits offered by such treatments, mortality from cardiovascular
disease remains high and significant unmet needs in the treatment
of cardiovascular disease remain.
As noted above, induction of HO-1 has been shown to be beneficial
in a variety of models of cardiovascular disease, and low levels of
HO-1 expression have been clinically correlated with elevated risk
of CV disease. Compounds of the invention, therefore, may be used
in treating or preventing a variety of cardiovascular disorders
including but not limited to atherosclerosis, hypertension,
myocardial infarction, chronic heart failure, stroke, subarachnoid
hemorrhage, and restenosis.
F. Diabetes
The compounds and methods of this invention may be used for
treating patients with diabetes. See U.S. patent application Ser.
No. 12/352,473, which is incorporated by reference herein in its
entirety. Diabetes is a complex disease characterized by the body's
failure to regulate circulating levels of glucose. This failure may
result from a lack of insulin, a peptide hormone that regulates
both the production and absorption of glucose in various tissues.
Deficient insulin compromises the ability of muscle, fat, and other
tissues to absorb glucose properly, leading to hyperglycemia
(abnormally high levels of glucose in the blood). Most commonly,
such insulin deficiency results from inadequate production in the
islet cells of the pancreas. In the majority of cases this arises
from autoimmune destruction of these cells, a condition known as
type 1 or juvenile-onset diabetes, but may also be due to physical
trauma or some other cause.
Diabetes may also arise when muscle and fat cells become less
responsive to insulin and do not absorb glucose properly, resulting
in hyperglycemia. This phenomenon is known as insulin resistance,
and the resulting condition is known as Type 2 diabetes. Type 2
diabetes, the most common type, is highly associated with obesity
and hypertension. Obesity is associated with an inflammatory state
of adipose tissue that is thought to play a major role in the
development of insulin resistance (e.g., Hotamisligil, 2006;
Guilherme et al., 2008).
Diabetes is associated with damage to many tissues, largely because
hyperglycemia (and hypoglycemia, which can result from excessive or
poorly timed doses of insulin) is a significant source of oxidative
stress. Chronic kidney failure, retinopathy, peripheral neuropathy,
peripheral vasculitis, and the development of dermal ulcers that
heal slowly or not at all are among the common complications of
diabetes. Because of their ability to protect against oxidative
stress, particularly by the induction of HO-1 expression, compounds
of the invention may be used in treatments for many complications
of diabetes. As noted above (Cai et al., 2005), chronic
inflammation and oxidative stress in the liver are suspected to be
primary contributing factors in the development of Type 2 diabetes.
Furthermore, PPAR.gamma. agonists such as thiazolidinediones are
capable of reducing insulin resistance and are known to be
effective treatments for Type 2 diabetes.
The effect of treatment of diabetes may be evaluated as follows.
Both the biological efficacy of the treatment modality as well as
the clinical efficacy are evaluated, if possible. For example,
because the disease manifests itself by increased blood sugar, the
biological efficacy of the treatment therefore can be evaluated,
for example, by observation of return of the evaluated blood
glucose towards normal. Measurement of glycosylated hemoglobin,
also called A1c or HbA1c, is another commonly used parameter of
blood glucose control. Measuring a clinical endpoint which can give
an indication of b-cell regeneration after, for example, a
six-month period of time, can give an indication of the clinical
efficacy of the treatment regimen.
G. Rheumatoid Arthritis
The compounds and methods of this invention may be used for
treating patients with RA. Typically the first signs of rheumatoid
arthritis (RA) appear in the synovial lining layer, with
proliferation of synovial fibroblasts and their attachment to the
articular surface at the joint margin (Lipsky, 1998). Subsequently,
macrophages, T cells and other inflammatory cells are recruited
into the joint, where they produce a number of mediators, including
the cytokines interleukin-1 (IL-1), which contributes to the
chronic sequelae leading to bone and cartilage destruction, and
tumour necrosis factor (TNF-.alpha.), which plays a role in
inflammation (Dinarello, 1998; Arend and Dayer, 1995; van den Berg,
2001). The concentration of IL-1 in plasma is significantly higher
in patients with RA than in healthy individuals and, notably,
plasma IL-1 levels correlate with RA disease activity (Eastgate et
al., 1988). Moreover, synovial fluid levels of IL-1 are correlated
with various radiographic and histologic features of RA (Kahle et
al., 1992; Rooney et al., 1990).
In normal joints, the effects of these and other proinflammatory
cytokines are balanced by a variety of anti-inflammatory cytokines
and regulatory factors (Burger and Dayer, 1995). The significance
of this cytokine balance is illustrated in juvenile RA patients,
who have cyclical increases in fever throughout the day (Prieur et
al., 1987). After each peak in fever, a factor that blocks the
effects of IL-1 is found in serum and urine. This factor has been
isolated, cloned and identified as IL-1 receptor antagonist
(IL-1ra), a member of the IL-1 gene family (Hannum et al., 1990).
IL-1ra, as its name indicates, is a natural receptor antagonist
that competes with IL-1 for binding to type I IL-1 receptors and,
as a result, blocks the effects of IL-1 (Arend et al., 1998). A 10-
to 100-fold excess of IL-1ra may be needed to block IL-1
effectively; however, synovial cells isolated from patients with RA
do not appear to produce enough IL-1ra to counteract the effects of
IL-1 (Firestein et al., 1994; Fujikawa et al., 1995).
H. Psoriatic Arthritis
The compounds and methods of this invention may be used for
treating patients with psoriatic arthritis. Psoriasis is an
inflammatory and proliferative skin disorder with a prevalence of
1.5-3%. Approximately 20% of patients with psoriasis develop a
characteristic form of arthritis that has several patterns
(Gladman, 1992; Jones et al., 1994; Gladman et al., 1995). Some
individuals present with joint symptoms first but in the majority,
skin psoriasis presents first. About one-third of patients have
simultaneous exacerbations of their skin and joint disease (Gladman
et al., 1987) and there is a topographic relationship between nail
and distal interphalangeal joint disease (Jones et al., 1994;
Wright, 1956). Although the inflammatory processes which link skin,
nail and joint disease remain elusive, an immune-mediated pathology
is implicated.
Psoriatic arthritis (PsA) is a chronic inflammatory arthropathy
characterized by the association of arthritis and psoriasis and was
recognized as a clinical entity distinct from rheumatoid arthritis
(RA) in 1964 (Blumberg et al., 1964). Subsequent studies have
revealed that PsA shares a number of genetic, pathogenic and
clinical features with other spondyloarthropathies (SpAs), a group
of diseases that comprise ankylosing spondylitis, reactive
arthritis and enteropathic arthritis (Wright, 1979). The notion
that PsA belongs to the SpA group has recently gained further
support from imaging studies demonstrating widespread enthesitis in
the, including PsA but not RA (McGonagle et al., 1999; McGonagle et
al., 1998). More specifically, enthesitis has been postulated to be
one of the earliest events occurring in the SpAs, leading to bone
remodeling and ankylosis in the spine, as well as to articular
synovitis when the inflamed entheses are close to peripheral
joints. However, the link between enthesitis and the clinical
manifestations in PsA remains largely unclear, as PsA can present
with fairly heterogeneous patterns of joint involvement with
variable degrees of severity (Marsal et al., 1999; Salvarani et
al., 1998). Thus, other factors must be posited to account for the
multifarious features of PsA, only a few of which (such as the
expression of the HLA-B27 molecule, which is strongly associated
with axial disease) have been identified. As a consequence, it
remains difficult to map the disease manifestations to specific
pathogenic mechanisms, which means that the treatment of this
condition remains largely empirical.
Family studies have suggested a genetic contribution to the
development of PsA (Moll and Wright, 1973). Other chronic
inflammatory forms of arthritis, such as ankylosing spondylitis and
rheumatoid arthritis, are thought to have a complex genetic basis.
However, the genetic component of PsA has been difficult to assess
for several reasons. There is strong evidence for a genetic
predisposition to psoriasis alone that may mask the genetic factors
that are important for the development of PsA. Although most would
accept PsA as a distinct disease entity, at times there is a
phenotypic overlap with rheumatoid arthritis and ankylosing
spondylitis. Also, PsA itself is not a homogeneous condition and
various subgroups have been proposed.
Increased amounts of TNF-.alpha. have been reported in both
psoriatic skin (Ettehadi et al., 1994) and synovial fluid (Partsch
et al., 1997). Recent trials have shown a positive benefit of
anti-TNF treatment in both PsA (Mease et al., 2000) and ankylosing
spondylitis (Brandt et al., 2000).
I. Reactive Arthritis
The compounds and methods of this invention may be used for
treating patients with reactive arthritis. In reactive arthritis
(ReA) the mechanism of joint damage is unclear, but it is likely
that cytokines play critical roles. A more prevalent Th1 profile
high levels of interferon gamma (IFN-.gamma.) and low levels of
interleukin 4 (IL-4) has been reported (Lahesmaa et al., 1992;
Schlaak et al., 1992; Simon et al., 1993; Schlaak et al., 1996;
Kotake et al., 1999; Ribbens et al., 2000), but several studies
have shown relative predominance of IL-4 and IL-10 and relative
lack of IFN-.gamma. and tumour necrosis factor alpha (TNF-.alpha.)
in the synovial membrane (Simon et al., 1994; Yin et al., 1999) and
fluid (SF) (Yin et al., 1999; Yin et al., 1997) of reactive
arthritis patients compared with rheumatoid arthritis (RA)
patients. A lower level of TNF-.alpha. secretion in reactive
arthritis than in RA patients has also been reported after ex vivo
stimulation of peripheral blood mononuclear cells (PBMC) (Braun et
al., 1999).
It has been argued that clearance of reactive arthritis-associated
bacteria requires the production of appropriate levels of
IFN-.gamma. and TNF-.alpha., while IL-10 acts by suppressing these
responses (Autenrieth et al., 1994; Sieper and Braun, 1995). IL-10
is a regulatory cytokine that inhibits the synthesis of IL-12 and
TNF-.gamma. by activated macrophages (de Waal et al., 1991; Hart et
al., 1995; Chomarat et al., 1995) and of IFN-.gamma. by T cells
(Macatonia et al., 1993).
J. Enteropathic Arthritis
The compounds and methods of this invention may be used for
treating patients with enteropathic arthritis. Typically
enteropathic arthritis (EA) occurs in combination with inflammatory
bowel diseases (IBD) such as Crohn's disease or ulcerative colitis.
It also can affect the spine and sacroiliac joints. Enteropathic
arthritis involves the peripheral joints, usually in the lower
extremities such as the knees or ankles. It commonly involves only
a few or a limited number of joints and may closely follow the
bowel condition. This occurs in approximately 11% of patients with
ulcerative colitis and 21% of those with Crohn's disease. The
synovitis is generally self-limited and non-deforming.
Enteropathic arthropathies comprise a collection of rheumatologic
conditions that share a link to GI pathology. These conditions
include reactive (i.e., infection-related) arthritis due to
bacteria (e.g., Shigella, Salmonella, Campylobacter, Yersinia
species, Clostridium difficile), parasites (e.g., Strongyloides
stercoralis, Taenia saginata, Giardia lamblia, Ascaris
lumbricoides, Cryptosporidium species), and spondyloarthropathies
associated with inflammatory bowel disease (IBD). Other conditions
and disorders include intestinal bypass (jejunoileal), arthritis,
celiac disease, Whipple disease, and collagenous colitis.
K. Juvenile Rheumatoid Arthritis
The compounds and methods of this invention may be used for
treating patients with JRA. Juvenile rheumatoid arthritis (JRA), a
term for the most prevalent form of arthritis in children, is
applied to a family of illnesses characterized by chronic
inflammation and hypertrophy of the synovial membranes. The term
overlaps, but is not completely synonymous, with the family of
illnesses referred to as juvenile chronic arthritis and/or juvenile
idiopathic arthritis in Europe.
Both innate and adaptive immune systems use multiple cell types, a
vast array of cell surface and secreted proteins, and
interconnected networks of positive and negative feedback (Lo et
al., 1999). Furthermore, while separable in thought, the innate and
adaptive wings of the immune system are functionally intersected
(Fearon and Locksley, 1996), and pathologic events occurring at
these intersecting points are likely to be highly relevant to our
understanding of pathogenesis of adult and childhood forms of
chronic arthritis (Warrington, et al., 2001).
Polyarticular JRA is a distinct clinical subtype characterized by
inflammation and synovial proliferation in multiple joints (four or
more), including the small joints of the hands (Jarvis, 2002). This
subtype of JRA may be severe, because of both its multiple joint
involvement and its capacity to progress rapidly over time.
Although clinically distinct, polyarticular JRA is not homogeneous,
and patients vary in disease manifestations, age of onset,
prognosis, and therapeutic response. These differences very likely
reflect a spectrum of variation in the nature of the immune and
inflammatory attack that can occur in this disease (Jarvis,
1998).
L. Early Inflammatory Arthritis
The compounds and methods of this invention may be used for
treating patients with early inflammatory arthritis. The clinical
presentation of different inflammatory arthropathies is similar
early in the course of disease. As a result, it is often difficult
to distinguish patients who are at risk of developing the severe
and persistent synovitis that leads to erosive joint damage from
those whose arthritis is more self-limited. Such distinction is
critical in order to target therapy appropriately, treating
aggressively those with erosive disease and avoiding unnecessary
toxicity in patients with more self-limited disease. Current
clinical criteria for diagnosing erosive arthropathies such as
rheumatoid arthritis (RA) are less effective in early disease and
traditional markers of disease activity such as joint counts and
acute phase response do not adequately identify patients likely to
have poor outcomes (Harrison et al., 1998). Parameters reflective
of the pathologic events occurring in the synovium are most likely
to be of significant prognostic value.
Recent efforts to identify predictors of poor outcome in early
inflammatory arthritis have identified the presence of RA specific
autoantibodies, in particular antibodies towards citrullinated
peptides, to be associated with erosive and persistent disease in
early inflammatory arthritis cohorts. On the basis of this, a
cyclical citrullinated peptide (CCP) has been developed to assist
in the identification of anti-CCP antibodies in patient sera. Using
this approach, the presence of anti-CCP antibodies has been shown
to be specific and sensitive for RA, can distinguish RA from other
arthropathies, and can potentially predict persistent, erosive
synovitis before these outcomes become clinically manifest.
Importantly, anti-CCP antibodies are often detectable in sera many
years prior to clinical symptoms suggesting that they may be
reflective of subclinical immune events (Nielen et al., 2004;
Rantapaa-Dahlqvist et al., 2003).
M. Ankylosing Spondylitis
The compounds and methods of this invention may be used for
treating patients with ankylosing spondylitis. AS is a disease
subset within a broader disease classification of
spondyloarthropathy. Patients affected with the various subsets of
spondyloarthropathy have disease etiologies that are often very
different, ranging from bacterial infections to inheritance. Yet,
in all subgroups, the end result of the disease process is axial
arthritis. Despite the early clinically differences seen in the
various patient populations, many of them end up nearly identical
after a disease course of ten-to-twenty years. Recent studies
suggest the mean time to clinical diagnosis of ankylosing
spondylitis from disease onset of disease is 7.5 years (Khan,
1998). These same studies suggest that the spondyloarthropathies
may have prevalence close to that of rheumatoid arthritis
(Feldtkeller et al., 2003; Doran et al., 2003).
AS is a chronic systemic inflammatory rheumatic disorder of the
axial skeleton with or without extraskeletal manifestations.
Sacroiliac joints and the spine are primarily affected, but hip and
shoulder joints, and less commonly peripheral joints or certain
extra-articular structures such as the eye, vasculature, nervous
system, and gastrointestinal system may also be involved. Its
etiology is not yet fully understood (Wordsworth, 1995; Calin and
Taurog, 1998). It is strongly associated with the major
histocompatibility class I (MHC I) HLA-B27 allele (Calin and
Taurog, 1998). AS affects individuals in the prime of their life
and is feared because of its potential to cause chronic pain and
irreversible damage of tendons, ligaments, joints, and bones
(Brewerton et al., 1973a; Brewerton et al., 1973b; Schlosstein et
al., 1973). AS may occur alone or in association with another form
of spondyloarthropathy such as reactive arthritis, psoriasis,
psoriatic arthritis, enthesitis, ulcerative colitis, irritable
bowel disease, or Crohn's disease, in which case it is classified
as secondary AS.
Typically, the affected sites include the discovertebral,
apophyseal, costovertebral, and costotransverse joints of the
spine, and the paravertebral ligamentous structures. Inflammation
of the entheses, which are sites of musculotendinous and
ligamentous attachment to bones, is also prominent in this disease
(Calin and Taurog, 1998). The site of enthesitis is known to be
infiltrated by plasma cells, lymphocytes, and polymorphonuclear
cells. The inflammatory process frequently results in gradual
fibrous and bony ankylosis, (Ball, 1971; Khan, 1990).
Delayed diagnosis is common because symptoms are often attributed
to more common back problems. A dramatic loss of flexibility in the
lumbar spine is an early sign of AS. Other common symptoms include
chronic pain and stiffness in the lower back which usually starts
where the lower spine is joined to the pelvis, or hip. Although
most symptoms begin in the lumbar and sacroiliac areas, they may
involve the neck and upper back as well. Arthritis may also occur
in the shoulder, hips and feet. Some patients have eye
inflammation, and more severe cases must be observed for heart
valve involvement.
The most frequent presentation is back pain, but disease can begin
atypically in peripheral joints, especially in children and women,
and rarely with acute iritis (anterior uveitis). Additional early
symptoms and signs are diminished chest expansion from diffuse
costovertebral involvement, low-grade fever, fatigue, anorexia,
weight loss, and anemia. Recurrent back pain--often nocturnal and
of varying intensity--is an eventual complaint, as is morning
stiffness typically relieved by activity. A flexed or bent-over
posture eases back pain and paraspinal muscle spasm; thus, some
degree of kyphosis is common in untreated patients.
Systemic manifestations occur in 1/3 of patients. Recurrent,
usually self-limited, acute iritis (anterior uveitis) rarely is
protracted and severe enough to impair vision. Neurologic signs can
occasionally result from compression radiculitis or sciatica,
vertebral fracture or subluxation, and cauda equina syndrome (which
consists of impotence, nocturnal urinary incontinence, diminished
bladder and rectal sensation, and absence of ankle jerks).
Cardiovascular manifestations can include aortic insufficiency,
angina, pericarditis, and ECG conduction abnormalities. A rare
pulmonary finding is upper lobe fibrosis, occasionally with
cavitation that may be mistaken for TB and can be complicated by
infection with Aspergillus.
AS is characterized by mild or moderate flares of active
spondylitis alternating with periods of almost or totally inactive
inflammation. Proper treatment in most patients results in minimal
or no disability and in full, productive lives despite back
stiffness. Occasionally, the course is severe and progressive,
resulting in pronounced incapacitating deformities. The prognosis
is bleak for patients with refractory iritis and for the rare
patient with secondary amyloidosis.
N. Ulcerative Colitis
The compounds and methods of this invention may be used for
treating patients with ulcerative colitis. Ulcerative colitis is a
disease that causes inflammation and sores, called ulcers, in the
lining of the large intestine. The inflammation usually occurs in
the rectum and lower part of the colon, but it may affect the
entire colon. Ulcerative colitis rarely affects the small intestine
except for the end section, called the terminal ileum. Ulcerative
colitis may also be called colitis or proctitis. The inflammation
makes the colon empty frequently, causing diarrhea. Ulcers form in
places where the inflammation has killed the cells lining the
colon; the ulcers bleed and produce pus.
Ulcerative colitis is an inflammatory bowel disease (IBD), the
general name for diseases that cause inflammation in the small
intestine and colon. Ulcerative colitis can be difficult to
diagnose because its symptoms are similar to other intestinal
disorders and to another type of IBD, Crohn's disease. Crohn's
disease differs from ulcerative colitis because it causes
inflammation deeper within the intestinal wall. Also, Crohn's
disease usually occurs in the small intestine, although it can also
occur in the mouth, esophagus, stomach, duodenum, large intestine,
appendix, and anus.
Ulcerative colitis may occur in people of any age, but most often
it starts between ages 15 and 30, or less frequently between ages
50 and 70. Children and adolescents sometimes develop the disease.
Ulcerative colitis affects men and women equally and appears to run
in some families. Theories about what causes ulcerative colitis
abound, but none have been proven. The most popular theory is that
the body's immune system reacts to a virus or a bacterium by
causing ongoing inflammation in the intestinal wall. People with
ulcerative colitis have abnormalities of the immune system, but
doctors do not know whether these abnormalities are a cause or a
result of the disease. Ulcerative colitis is not caused by
emotional distress or sensitivity to certain foods or food
products, but these factors may trigger symptoms in some
people.
The most common symptoms of ulcerative colitis are abdominal pain
and bloody diarrhea. Patients also may experience fatigue, weight
loss, loss of appetite, rectal bleeding, and loss of body fluids
and nutrients. About half of patients have mild symptoms. Others
suffer frequent fever, bloody diarrhea, nausea, and severe
abdominal cramps. Ulcerative colitis may also cause problems such
as arthritis, inflammation of the eye, liver disease (hepatitis,
cirrhosis, and primary sclerosing cholangitis), osteoporosis, skin
rashes, and anemia. No one knows for sure why problems occur
outside the colon. Scientists think these complications may occur
when the immune system triggers inflammation in other parts of the
body. Some of these problems go away when the colitis is
treated.
A thorough physical exam and a series of tests may be required to
diagnose ulcerative colitis. Blood tests may be done to check for
anemia, which could indicate bleeding in the colon or rectum. Blood
tests may also uncover a high white blood cell count, which is a
sign of inflammation somewhere in the body. By testing a stool
sample, the doctor can detect bleeding or infection in the colon or
rectum. The doctor may do a colonoscopy or sigmoidoscopy. For
either test, the doctor inserts an endoscope--a long, flexible,
lighted tube connected to a computer and TV monitor--into the anus
to see the inside of the colon and rectum. The doctor will be able
to see any inflammation, bleeding, or ulcers on the colon wall.
During the exam, the doctor may do a biopsy, which involves taking
a sample of tissue from the lining of the colon to view with a
microscope. A barium enema x ray of the colon may also be required.
This procedure involves filling the colon with barium, a chalky
white solution. The barium shows up white on x-ray film, allowing
the doctor a clear view of the colon, including any ulcers or other
abnormalities that might be there.
Treatment for ulcerative colitis depends on the seriousness of the
disease. Most people are treated with medication. In severe cases,
a patient may need surgery to remove the diseased colon. Surgery is
the only cure for ulcerative colitis. Some people whose symptoms
are triggered by certain foods are able to control the symptoms by
avoiding foods that upset their intestines, like highly seasoned
foods, raw fruits and vegetables, or milk sugar (lactose). Each
person may experience ulcerative colitis differently, so treatment
is adjusted for each individual. Emotional and psychological
support is important. Some people have remissions--periods when the
symptoms go away--that last for months or even years. However, most
patients' symptoms eventually return. This changing pattern of the
disease means one cannot always tell when a treatment has helped.
Some people with ulcerative colitis may need medical care for some
time, with regular doctor visits to monitor the condition.
O. Crohn's Disease
The compounds and methods of this invention may be used for
treating patients with Crohn's disease. Another disorder for which
immunosuppression has been tried is Crohn's disease. Crohn's
disease symptoms include intestinal inflammation and the
development of intestinal stenosis and fistulas; neuropathy often
accompanies these symptoms. Anti-inflammatory drugs, such as
5-aminosalicylates (e.g., mesalamine) or corticosteroids, are
typically prescribed, but are not always effective (reviewed in
Botoman et al., 1998). Immunosuppression with cyclosporine is
sometimes beneficial for patients resistant to or intolerant of
corticosteroids (Brynskov et al., 1989).
Efforts to develop diagnostic and treatment tools against Crohn's
disease have focused on the central role of cytokines (Schreiber,
1998; van Hogezand and Verspaget, 1998). Cytokines are small
secreted proteins or factors (5 to 20 kD) that have specific
effects on cell-to-cell interactions, intercellular communication,
or the behavior of other cells. Cytokines are produced by
lymphocytes, especially T.sub.H1 and T.sub.H2 lymphocytes,
monocytes, intestinal macrophages, granulocytes, epithelial cells,
and fibroblasts (reviewed in Rogler and Andus, 1998; Galley and
Webster, 1996). Some cytokines are pro-inflammatory (e.g.,
TNF-.alpha., IL-1 (.alpha. and .beta.), IL-6, IL-8, IL-12, or
leukemia inhibitory factor [LIF]); others are anti-inflammatory
(e.g., IL-1 receptor antagonist, IL-4, IL-10, IL-11, and
TGF-.beta.). However, there may be overlap and functional
redundancy in their effects under certain inflammatory
conditions.
In active cases of Crohn's disease, elevated concentrations of
TNF-.alpha. and IL-6 are secreted into the blood circulation, and
TNF-.alpha., IL-1, IL-6, and IL-8 are produced in excess locally by
mucosal cells (id.; Funakoshi et al., 1998). These cytokines can
have far-ranging effects on physiological systems including bone
development, hematopoiesis, and liver, thyroid, and
neuropsychiatric function. Also, an imbalance of the
IL-1.beta./IL-1ra ratio, in favor of pro-inflammatory IL-1.beta.,
has been observed in patients with Crohn's disease (Rogler and
Andus, 1998; Saiki et al., 1998; Dionne et al., 1998; but see
Kuboyama, 1998). One study suggested that cytokine profiles in
stool samples could be a useful diagnostic tool for Crohn's disease
(Saiki et al., 1998).
Treatments that have been proposed for Crohn's disease include the
use of various cytokine antagonists (e.g., IL-1ra), inhibitors
(e.g., of IL-1.beta. converting enzyme and antioxidants) and
anti-cytokine antibodies (Rogler and Andus, 1998; van Hogezand and
Verspaget, 1998; Reimund et al., 1998; Lugering et al., 1998;
McAlindon et al., 1998). In particular, monoclonal antibodies
against TNF-.alpha. have been tried with some success in the
treatment of Crohn's disease (Targan et al., 1997; Stack et al.,
1997; van Dullemen et al., 1995). These compounds may be used in
combination therapy with compounds of the present disclosure.
Another approach to the treatment of Crohn's disease has focused on
at least partially eradicating the bacterial community that may be
triggering the inflammatory response and replacing it with a
non-pathogenic community. For example, U.S. Pat. No. 5,599,795
discloses a method for the prevention and treatment of Crohn's
disease in human patients. Their method was directed to sterilizing
the intestinal tract with at least one antibiotic and at least one
anti-fungal agent to kill off the existing flora and replacing them
with different, select, well-characterized bacteria taken from
normal humans. Borody taught a method of treating Crohn's disease
by at least partial removal of the existing intestinal microflora
by lavage and replacement with a new bacterial community introduced
by fecal inoculum from a disease-screened human donor or by a
composition comprising Bacteroides and Escherichia coli species.
(U.S. Pat. No. 5,443,826).
P. Systemic Lupus Erythematosus
The compounds and methods of this invention may be used for
treating patients with SLE. There has also been no known cause for
autoimmune diseases such as systemic lupus erythematosus. Systemic
lupus erythematosus (SLE) is an autoimmune rheumatic disease
characterized by deposition in tissues of autoantibodies and immune
complexes leading to tissue injury (Kotzin, 1996). In contrast to
autoimmune diseases such as MS and type 1 diabetes mellitus, SLE
potentially involves multiple organ systems directly, and its
clinical manifestations are diverse and variable (reviewed by
Kotzin and O'Dell, 1995). For example, some patients may
demonstrate primarily skin rash and joint pain, show spontaneous
remissions, and require little medication. At the other end of the
spectrum are patients who demonstrate severe and progressive kidney
involvement that requires therapy with high doses of steroids and
cytotoxic drugs such as cyclophosphamide (Kotzin, 1996).
The serological hallmark of SLE, and the primary diagnostic test
available, is elevated serum levels of IgG antibodies to
constituents of the cell nucleus, such as double-stranded DNA
(dsDNA), single-stranded DNA (ss-DNA), and chromatin. Among these
autoantibodies, IgG anti-dsDNA antibodies play a major role in the
development of lupus glomerulonephritis (G N) (Hahn and Tsao, 1993;
Ohnishi et al., 1994). Glomerulonephritis is a serious condition in
which the capillary walls of the kidney's blood purifying glomeruli
become thickened by accretions on the epithelial side of glomerular
basement membranes. The disease is often chronic and progressive
and may lead to eventual renal failure.
Q. Irritable Bowel Syndrome
The compounds and methods of this invention may be used for
treating patients with Irritable bowel syndrome (IBS). IBS is a
functional disorder characterized by abdominal pain and altered
bowel habits. This syndrome may begin in young adulthood and can be
associated with significant disability. This syndrome is not a
homogeneous disorder. Rather, subtypes of IBS have been described
on the basis of the predominant symptom--diarrhea, constipation, or
pain. In the absence of "alarm" symptoms, such as fever, weight
loss, and gastrointestinal bleeding, a limited workup is needed.
Once a diagnosis of IBS is made, an integrated treatment approach
can effectively reduce the severity of symptoms. IBS is a common
disorder, although its prevalence rates have varied. In general,
IBS affects about 15% of US adults and occurs about three times
more often in women than in men (Jailwala et al., 2000).
IBS accounts for between 2.4 million and 3.5 million visits to
physicians each year. It not only is the most common condition seen
by gastroenterologists but also is one of the most common
gastrointestinal conditions seen by primary care physicians
(Everhart et al., 1991; Sandler, 1990).
IBS is also a costly disorder. Compared with persons who do not
have bowel symptoms, persons with IBS miss three times as many
workdays and are more likely to report being too sick to work
(Drossman et al., 1993; Drossman et al., 1997). Moreover, those
with IBS incur hundreds of dollars more in medical charges than
persons without bowel disorders (Talley et al., 1995).
No specific abnormality accounts for the exacerbations and
remissions of abdominal pain and altered bowel habits experienced
by patients with IBS. The evolving theory of IBS suggests
dysregulation at multiple levels of the brain-gut axis.
Dysmotility, visceral hypersensitivity, abnormal modulation of the
central nervous system (CNS), and infection have all been
implicated. In addition, psychosocial factors play an important
modifying role. Abnormal intestinal motility has long been
considered a factor in the pathogenesis of IBS. Transit time
through the small intestine after a meal has been shown to be
shorter in patients with diarrhea-predominant IBS than in patients
who have the constipation-predominant or pain-predominant subtype
(Cann et al., 1983).
In studies of the small intestine during fasting, the presence of
both discrete, clustered contractions and prolonged, propagated
contractions has been reported in patients with IBS (Kellow and
Phillips, 1987). They also experience pain with irregular
contractions more often than healthy persons (Kellow and Phillips,
1987; Horwitz and Fisher, 2001)
These motility findings do not account for the entire symptom
complex in patients with IBS; in fact, most of these patients do
not have demonstrable abnormalities (Rothstein, 2000). Patients
with IBS have increased sensitivity to visceral pain. Studies
involving balloon distention of the rectosigmoid colon have shown
that patients with IBS experience pain and bloating at pressures
and volumes much lower than control subjects (Whitehead et al.,
1990). These patients maintain normal perception of somatic
stimuli.
Multiple theories have been proposed to explain this phenomenon.
For example, receptors in the viscera may have increased
sensitivity in response to distention or intraluminal contents.
Neurons in the dorsal horn of the spinal cord may have increased
excitability. In addition, alteration in CNS processing of
sensations may be involved (Drossman et al., 1997). Functional
magnetic resonance imaging studies have recently shown that
compared with control subjects, patients with IBS have increased
activation of the anterior cingulate cortex, an important pain
center, in response to a painful rectal stimulus (Mertz et al.,
2000).
Increasingly, evidence suggests a relationship between infectious
enteritis and subsequent development of IBS. Inflammatory cytokines
may play a role. In a survey of patients with a history of
confirmed bacterial gastroenteritis (Neal et al., 1997), 25%
reported persistent alteration of bowel habits. Persistence of
symptoms may be due to psychological stress at the time of acute
infection (Gwee et al., 1999).
Recent data suggest that bacterial overgrowth in the small
intestine may have a role in IBS symptoms. In one study (Pimentel
et al., 2000), 157 (78%) of 202 IBS patients referred for hydrogen
breath testing had test findings that were positive for bacterial
overgrowth. Of the 47 subjects who had follow-up testing, 25 (53%)
reported improvement in symptoms (i.e., abdominal pain and
diarrhea) with antibiotic treatment.
IBS may present with a range of symptoms. However, abdominal pain
and altered bowel habits remain the primary features. Abdominal
discomfort is often described as crampy in nature and located in
the left lower quadrant, although the severity and location can
differ greatly. Patients may report diarrhea, constipation, or
alternating episodes of diarrhea and constipation. Diarrheal
symptoms are typically described as small-volume, loose stools, and
stool is sometimes accompanied by mucus discharge. Patients also
may report bloating, fecal urgency, incomplete evacuation, and
abdominal distention. Upper gastrointestinal symptoms, such as
gastroesophageal reflux, dyspepsia, or nausea, may also be present
(Lynn and Friedman, 1993).
Persistence of symptoms is not an indication for further testing;
it is a characteristic of IBS and is itself an expected symptom of
the syndrome. More extensive diagnostic evaluation is indicated in
patients whose symptoms are worsening or changing. Indications for
further testing also include presence of alarm symptoms, onset of
symptoms after age 50, and a family history of colon cancer. Tests
may include colonoscopy, computed tomography of the abdomen and
pelvis, and barium studies of the small or large intestine.
R. Sjogren's Syndrome
The compounds and methods of this invention may be used for
treating patients with SS. Primary Sjogren's syndrome (SS) is a
chronic, slowly progressive, systemic autoimmune disease, which
affects predominantly middle-aged women (female-to-male ratio 9:1),
although it can be seen in all ages including childhood (Jonsson et
al., 2002). It is characterized by lymphocytic infiltration and
destruction of the exocrine glands, which are infiltrated by
mononuclear cells including CD4+, CD8+ lymphocytes and B-cells
(Jonsson et al., 2002). In addition, extraglandular (systemic)
manifestations are seen in one-third of patients (Jonsson et al.,
2001).
The glandular lymphocytic infiltration is a progressive feature
(Jonsson et al., 1993), which, when extensive, may replace large
portions of the organs. Interestingly, the glandular infiltrates in
some patients closely resemble ectopic lymphoid microstructures in
the salivary glands (denoted as ectopic germinal centers)
(Salomonsson et al., 2002; Xanthou et al., 2001). In SS, ectopic
GCs are defined as T and B cell aggregates of proliferating cells
with a network of follicular dendritic cells and activated
endothelial cells. These GC-like structures formed within the
target tissue also portray functional properties with production of
autoantibodies (anti-Ro/SSA and anti-La/SSB) (Salomonsson and
Jonsson, 2003).
In other systemic autoimmune diseases, such as RA, factors critical
for ectopic GCs have been identified. Rheumatoid synovial tissues
with GCs were shown to produce chemokines CXCL13, CCL21 and
lymphotoxin (LT)-.beta. (detected on follicular center and mantle
zone B cells). Multivariate regression analysis of these analytes
identified CXCL13 and LT-.beta. as the solitary cytokines
predicting GCs in rheumatoid synovitis (Weyand and Goronzy, 2003).
Recently CXCL13 and CXCR5 in salivary glands has been shown to play
an essential role in the inflammatory process by recruiting B and T
cells, therefore contributing to lymphoid neogenesis and ectopic GC
formation in SS (Salomonsson et al., 2002).
S. Psoriasis
The compounds and methods of this invention may be used for
treating patients with psoriasis. Psoriasis is a chronic skin
disease of scaling and inflammation that affects 2 to 2.6 percent
of the United States population, or between 5.8 and 7.5 million
people. Although the disease occurs in all age groups, it primarily
affects adults. It appears about equally in males and females.
Psoriasis occurs when skin cells quickly rise from their origin
below the surface of the skin and pile up on the surface before
they have a chance to mature. Usually this movement (also called
turnover) takes about a month, but in psoriasis it may occur in
only a few days. In its typical form, psoriasis results in patches
of thick, red (inflamed) skin covered with silvery scales. These
patches, which are sometimes referred to as plaques, usually itch
or feel sore. They most often occur on the elbows, knees, other
parts of the legs, scalp, lower back, face, palms, and soles of the
feet, but they can occur on skin anywhere on the body. The disease
may also affect the fingernails, the toenails, and the soft tissues
of the genitals and inside the mouth. While it is not unusual for
the skin around affected joints to crack, approximately 1 million
people with psoriasis experience joint inflammation that produces
symptoms of arthritis. This condition is called psoriatic
arthritis.
Psoriasis is a skin disorder driven by the immune system,
especially involving a type of white blood cell called a T cell.
Normally, T cells help protect the body against infection and
disease. In the case of psoriasis, T cells are put into action by
mistake and become so active that they trigger other immune
responses, which lead to inflammation and to rapid turnover of skin
cells. In about one-third of the cases, there is a family history
of psoriasis. Researchers have studied a large number of families
affected by psoriasis and identified genes linked to the disease.
People with psoriasis may notice that there are times when their
skin worsens, then improves. Conditions that may cause flareups
include infections, stress, and changes in climate that dry the
skin. Also, certain medicines, including lithium and beta blockers,
which are prescribed for high blood pressure, may trigger an
outbreak or worsen the disease.
T. Infectious Diseases
Compounds of the present disclosure may be useful in the treatment
of infectious diseases, including viral and bacterial infections.
As noted above, such infections may be associated with severe
localized or systemic inflammatory responses. For example,
influenza may cause severe inflammation of the lung and bacterial
infection can cause the systemic hyperinflammatory response,
including the excessive production of multiple inflammatory
cytokines, that is the hallmark of sepsis. In addition, compounds
of the invention may be useful in directly inhibiting the
replication of viral pathogens. Previous studies have demonstrated
that related compounds such as CDDO can inhibit the replication of
HIV in macrophages (Vazquez et al., 2005). Other studies have
indicated that inhibition of NF-kappa B signaling may inhibit
influenza virus replication, and that cyclopentenone prostaglandins
may inhibit viral replication (e.g., Mazur et al., 2007; Pica et
al., 2000).
VI. PHARMACEUTICAL FORMULATIONS AND ROUTES OF ADMINISTRATION
The compounds of the present disclosure may be administered by a
variety of methods, e.g., orally or by injection (e.g.
subcutaneous, intravenous, intraperitoneal, etc.). Depending on the
route of administration, the active compounds may be coated in a
material to protect the compound from the action of acids and other
natural conditions which may inactivate the compound. They may also
be administered by continuous perfusion/infusion of a disease or
wound site.
To administer the therapeutic compound by other than parenteral
administration, it may be necessary to coat the compound with, or
co-administer the compound with, a material to prevent its
inactivation. For example, the therapeutic compound may be
administered to a patient in an appropriate carrier, for example,
liposomes, or a diluent. Pharmaceutically acceptable diluents
include saline and aqueous buffer solutions. Liposomes include
water-in-oil-in-water CGF emulsions as well as conventional
liposomes (Strejan et al., 1984).
The therapeutic compound may also be administered parenterally,
intraperitoneally, intraspinally, or intracerebrally. Dispersions
can be prepared in glycerol, liquid polyethylene glycols, and
mixtures thereof and in oils. Under ordinary conditions of storage
and use, these preparations may contain a preservative to prevent
the growth of microorganisms.
Pharmaceutical compositions suitable for injectable use include
sterile aqueous solutions (where water soluble) or dispersions and
sterile powders for the extemporaneous preparation of sterile
injectable solutions or dispersion. In all cases, the composition
must be sterile and must be fluid to the extent that easy
syringability exists. It must be stable under the conditions of
manufacture and storage and must be preserved against the
contaminating action of microorganisms such as bacteria and fungi.
The carrier can be a solvent or dispersion medium containing, for
example, water, ethanol, polyol (such as, glycerol, propylene
glycol, and liquid polyethylene glycol, and the like), suitable
mixtures thereof, and vegetable oils. The proper fluidity can be
maintained, for example, by the use of a coating such as lecithin,
by the maintenance of the required particle size in the case of
dispersion and by the use of surfactants. Prevention of the action
of microorganisms can be achieved by various antibacterial and
antifungal agents, for example, parabens, chlorobutanol, phenol,
ascorbic acid, thimerosal, and the like. In many cases, it will be
preferable to include isotonic agents, for example, sugars, sodium
chloride, or polyalcohols such as mannitol and sorbitol, in the
composition. Prolonged absorption of the injectable compositions
can be brought about by including in the composition an agent which
delays absorption, for example, aluminum monostearate or
gelatin.
Sterile injectable solutions can be prepared by incorporating the
therapeutic compound in the required amount in an appropriate
solvent with one or a combination of ingredients enumerated above,
as required, followed by filtered sterilization. Generally,
dispersions are prepared by incorporating the therapeutic compound
into a sterile carrier which contains a basic dispersion medium and
the required other ingredients from those enumerated above. In the
case of sterile powders for the preparation of sterile injectable
solutions, the preferred methods of preparation are vacuum drying
and freeze-drying which yields a powder of the active ingredient
(i.e., the therapeutic compound) plus any additional desired
ingredient from a previously sterile-filtered solution thereof.
The therapeutic compound can be orally administered, for example,
with an inert diluent or an assimilable edible carrier. The
therapeutic compound and other ingredients may also be enclosed in
a hard or soft shell gelatin capsule, compressed into tablets, or
incorporated directly into the subject's diet. For oral therapeutic
administration, the therapeutic compound may be incorporated with
excipients and used in the form of ingestible tablets, buccal
tablets, troches, capsules, elixirs, suspensions, syrups, wafers,
and the like. The percentage of the therapeutic compound in the
compositions and preparations may, of course, be varied. The amount
of the therapeutic compound in such therapeutically useful
compositions is such that a suitable dosage will be obtained.
It is especially advantageous to formulate parenteral compositions
in dosage unit form for ease of administration and uniformity of
dosage. Dosage unit form as used herein refers to physically
discrete units suited as unitary dosages for the subjects to be
treated; each unit containing a predetermined quantity of
therapeutic compound calculated to produce the desired therapeutic
effect in association with the required pharmaceutical carrier. The
specification for the dosage unit forms of the invention are
dictated by and directly dependent on (a) the unique
characteristics of the therapeutic compound and the particular
therapeutic effect to be achieved, and (b) the limitations inherent
in the art of compounding such a therapeutic compound for the
treatment of a selected condition in a patient.
The therapeutic compound may also be administered topically to the
skin, eye, or mucosa. Alternatively, if local delivery to the lungs
is desired the therapeutic compound may be administered by
inhalation in a dry-powder or aerosol formulation.
Active compounds are administered at a therapeutically effective
dosage sufficient to treat a condition associated with a condition
in a patient. A "therapeutically effective amount" preferably
reduces the amount of symptoms of the condition in the infected
patient by at least about 20%, more preferably by at least about
40%, even more preferably by at least about 60%, and still more
preferably by at least about 80% relative to untreated subjects.
For example, the efficacy of a compound can be evaluated in an
animal model system that may be predictive of efficacy in treating
the disease in humans, such as the model systems shown in the
examples and drawings.
The actual dosage amount of a compound of the present disclosure or
composition comprising a compound of the present disclosure
administered to a subject may be determined by physical and
physiological factors such as age, sex, body weight, severity of
condition, the type of disease being treated, previous or
concurrent therapeutic interventions, idiopathy of the subject and
on the route of administration. These factors may be determined by
a skilled artisan. The practitioner responsible for administration
will typically determine the concentration of active ingredient(s)
in a composition and appropriate dose(s) for the individual
subject. The dosage may be adjusted by the individual physician in
the event of any complication.
An effective amount typically will vary from about 0.001 mg/kg to
about 1,000 mg/kg, from about 0.01 mg/kg to about 750 mg/kg, from
about 100 mg/kg to about 500 mg/kg, from about 1.0 mg/kg to about
250 mg/kg, from about 10.0 mg/kg to about 150 mg/kg in one or more
dose administrations daily, for one or several days (depending, of
course, of the mode of administration and the factors discussed
above). Other suitable dose ranges include 1 mg to 10,000 mg per
day, 100 mg to 10,000 mg per day, 500 mg to 10,000 mg per day, and
500 mg to 1,000 mg per day. In some particular embodiments, the
amount is less than 10,000 mg per day with a range, for example, of
750 mg to 9,000 mg per day.
The effective amount may be less than 1 mg/kg/day, less than 500
mg/kg/day, less than 250 mg/kg/day, less than 100 mg/kg/day, less
than 50 mg/kg/day, less than 25 mg/kg/day or less than 10
mg/kg/day. It may alternatively be in the range of 1 mg/kg/day to
200 mg/kg/day. For example, regarding treatment of diabetic
patients, the unit dosage may be an amount that reduces blood
glucose by at least 40% as compared to an untreated subject. In
another embodiment, the unit dosage is an amount that reduces blood
glucose to a level that is .+-.10% of the blood glucose level of a
non-diabetic subject.
In other non-limiting examples, a dose may also comprise from about
1 microgram/kg/body weight, about 5 microgram/kg/body weight, about
10 micro-gram/kg/body weight, about 50 microgram/kg/body weight,
about 100 micro-gram/kg/body weight, about 200 microgram/kg/body
weight, about 350 micro-gram/kg/body weight, about 500
microgram/kg/body weight, about 1 milligram/kg/body weight, about 5
milligram/kg/body weight, about 10 milligram/kg/body weight, about
50 milligram/kg/body weight, about 100 milligram/kg/body weight,
about 200 milli-gram/kg/body weight, about 350 milligram/kg/body
weight, about 500 milligram/kg/body weight, to about 1,000
mg/kg/body weight or more per administration, and any range
derivable therein. In non-limiting examples of a derivable range
from the numbers listed herein, a range of about 5 mg/kg/body
weight to about 100 mg/kg/body weight, about 5 microgram/kg/body
weight to about 500 milligram/kg/body weight, etc., can be
administered, based on the numbers described above.
In certain embodiments, a pharmaceutical composition of the present
disclosure may comprise, for example, at least about 0.1% of a
compound of the present disclosure.
In other embodiments, the compound of the present disclosure may
comprise between about 2% to about 75% of the weight of the unit,
or between about 25% to about 60%, for example, and any range
derivable therein.
Single or multiple doses of the agents are contemplated. Desired
time intervals for delivery of multiple doses can be determined by
one of ordinary skill in the art employing no more than routine
experimentation. As an example, subjects may be administered two
doses daily at approximately 12 hour intervals. In some
embodiments, the agent is administered once a day.
The agent(s) may be administered on a routine schedule. As used
herein a routine schedule refers to a predetermined designated
period of time. The routine schedule may encompass periods of time
which are identical or which differ in length, as long as the
schedule is predetermined. For instance, the routine schedule may
involve administration twice a day, every day, every two days,
every three days, every four days, every five days, every six days,
a weekly basis, a monthly basis or any set number of days or weeks
there-between. Alternatively, the predetermined routine schedule
may involve administration on a twice daily basis for the first
week, followed by a daily basis for several months, etc. In other
embodiments, the invention provides that the agent(s) may taken
orally and that the timing of which is or is not dependent upon
food intake. Thus, for example, the agent can be taken every
morning and/or every evening, regardless of when the subject has
eaten or will eat.
VII. COMBINATION THERAPY
In addition to being used as a monotherapy, the compounds of the
present disclosure may also find use in combination therapies.
Effective combination therapy may be achieved with a single
composition or pharmacological formulation that includes both
agents, or with two distinct compositions or formulations, at the
same time, wherein one composition includes a compound of this
invention, and the other includes the second agent(s).
Alternatively, the therapy may precede or follow the other agent
treatment by intervals ranging from minutes to months.
Various combinations may be employed, such as when a compound of
the present disclosure is "A" and "B" represents a secondary agent,
non-limiting examples of which are described below:
TABLE-US-00003 A/B/A B/A/B B/B/A A/A/B A/B/B B/A/A A/B/B/B B/A/B/B
B/B/B/A B/B/A/B A/A/B/B A/B/A/B A/B/B/A B/B/A/A B/A/B/A B/A/A/B
A/A/A/B B/A/A/A A/B/A/A A/A/B/A
Administration of the compounds of the present disclosure to a
patient will follow general protocols for the administration of
pharmaceuticals, taking into account the toxicity, if any, of the
drug. It is expected that the treatment cycles would be repeated as
necessary.
Beta interferons may be suitable secondary agents. These are
medications derived from human cytokines which help regulate the
immune system. They include interferon .beta.-1b and interferon
.beta.-1a. Betaseron has been approved by the FDA for relapsing
forms of secondary progressive MS. Furthermore, the FDA has
approved the use of several .beta.-interferons as treatments for
people who have experienced a single attack that suggests multiple
sclerosis, and who may be at risk of future attacks and developing
definite MS. For example, risk of MS may be suggested when an MRI
scan of the brain shows lesions that predict a high risk of
conversion to definite MS.
Glatiramer acetate is a further example of a secondary agent that
may be used in a combination treatment. Glatiramer is presently
used to treat relapsing remitting MS. It is made of four amino
acids that are found in myelin. This drug is reported to stimulate
T cells in the body's immune system to change from harmful,
pro-inflammatory agents to beneficial, anti-inflammatory agents
that work to reduce inflammation at lesion sites.
Another potential secondary agent is mitoxantrone, a chemotherapy
drug used for many cancers. This drug is also FDA-approved for
treatment of aggressive forms of relapsing remitting MS, as well as
certain forms of progressive MS. It is given intravenously,
typically every three months. This medication is effective, but is
limited by cardiac toxicity. Novantrone has been approved by the
FDA for secondary progressive, progressive-relapsing, and worsening
relapsing-remitting MS.
Another potential secondary agent is natalizumab. In general,
natalizumab works by blocking the attachment of immune cells to
brain blood vessels, which is a necessary step for immune cells to
cross into the brain, thus reducing the immune cells' inflammatory
action on brain neurons. Natalizumab has been shown to
significantly reduce the frequency of attacks in people with
relapsing MS.
In the case of relapsing remitting MS, patients may be given
intravenous corticosteroids, such as methylprednisolone, as a
secondary agent, to end the attack sooner and leave fewer lasting
deficits.
Other common drugs for MS that may be used in combination with the
present oleanolic acid derivatives include immunosuppressive drugs
such as azathioprine, cladribine and cyclophosphamide.
It is contemplated that other anti-inflammatory agents may be used
in conjunction with the treatments of the current invention. Other
COX inhibitors may be used, including arylcarboxylic acids
(salicylic acid, acetylsalicylic acid, diflunisal, choline
magnesium trisalicylate, salicylate, benorylate, flufenamic acid,
mefenamic acid, meclofenamic acid and triflumic acid), arylalkanoic
acids (diclofenac, fenclofenac, alclofenac, fentiazac, ibuprofen,
flurbiprofen, ketoprofen, naproxen, fenoprofen, fenbufen, suprofen,
indoprofen, tiaprofenic acid, benoxaprofen, pirprofen, tolmetin,
zomepirac, clopinac, indomethacin and sulindac) and enolic acids
(phenylbutazone, oxyphenbutazone, azapropazone, feprazone,
piroxicam, and isoxicam. See also U.S. Pat. No. 6,025,395, which is
incorporated herein by reference.
Histamine H2 receptor blocking agents may also be used in
conjunction with the compounds of the current invention, including
cimetidine, ranitidine, famotidine and nizatidine.
Treatment with acetylcholinesterase inhibitors such as tacrine,
donepizil, metrifonate and rivastigmine for the treatment of
Alzheimer's and other disease in conjunction with the compounds of
the present disclosure is contemplated. Other acetylcholinesterase
inhibitors may be developed which may be used once approved include
rivastigmine and metrifonate. Acetylcholinesterase inhibitors
increase the amount of neurotransmitter acetylcholine at the nerve
terminal by decreasing its breakdown by the enzyme
cholinesterase.
MAO-B inhibitors such as selegilene may be used in conjunction with
the compounds of the current invention. Selegilene is used for
Parkinson's disease and irreversibly inhibits monoamine oxidase
type B (MAO-B). Monoamine oxidase is an enzyme that inactivates the
monoamine neurotransmitters norepinephrine, serotonin and
dopamine.
Dietary and nutritional supplements with reported benefits for
treatment or prevention of Parkinson's, Alzheimer's, multiple
sclerosis, amyotrophic lateral sclerosis, rheumatoid arthritis,
inflammatory bowel disease, and all other diseases whose
pathogenesis is believed to involve excessive production of either
nitric oxide (NO) or prostaglandins, such as acetyl-L-carnitine,
octacosanol, evening primrose oil, vitamin B6, tyrosine,
phenylalanine, vitamin C, L-dopa, or a combination of several
antioxidants may be used in conjunction with the compounds of the
current invention.
For the treatment or prevention of cancer, compounds of the
invention may be combined with one or more of the following:
radiation, chemotherapy agents (e.g., cytotoxic agents such as
anthracyclines, vincristine, vinblastin, microtubule-targeting
agents such as paclitaxel and docetaxel, 5-FU and related agents,
cisplatin and other platinum-containing compounds, irinotecan and
topotecan, gemcitabine, temozolomide, etc.), targeted therapies
(e.g., imatinib, bortezomib, bevacizumab, rituximab), or vaccine
therapies designed to promote an enhanced immune response targeting
cancer cells.
For the treatment or prevention of autoimmune disease, compounds of
the invention may be combined with one or more of the following:
corticosteroids, methotrexate, anti-TNF antibodies, other
TNF-targeting protein therapies, and NSAIDs. For the treatment of
prevention of cardiovascular diseases, compounds of the invention
may be combined with antithrombotic therapies, anticholesterol
therapies such as statins (e.g., atorvastatin), and surgical
interventions such as stenting or coronary artery bypass grafting.
For the treatment of osteoporosis, compounds of the invention may
be combined with antiresorptive agents such as bisphosphonates or
anabolic therapies such as teriparatide or parathyroid hormone. For
the treatment of neuropsychiatric conditions, compounds of the
invention may be combined with antidepressants (e.g., imipramine or
SSRIs such as fluoxetine), antipsychotic agents (e.g., olanzapine,
sertindole, risperidone), mood stabilizers (e.g., lithium,
valproate semisodium), or other standard agents such as anxiolytic
agents. For the treatment of neurological disorders, compounds of
the invention may be combined with anticonvulsant agents (e.g.,
valproate semisodium, gabapentin, phenyloin, carbamazepine, and
topiramate), antithrombotic agents (e.g., tissue plasminogen
activator), or analgesics (e.g., opioids, sodium channel blockers,
and other antinociceptive agents).
For the treatment of disorders involving oxidative stress,
compounds of the present disclosure may be combined with
tetrahydrobiopterin (BH4) or related compounds. BH4 is a cofactor
for constitutive forms of nitric oxide synthase, and may be
depleted by reactions with peroxynitrite. Peroxynitrite is formed
by the reaction of nitric oxide and superoxide. Thus, under
conditions of oxidative stress excessive levels of superoxide can
deplete normal, beneficial levels of nitric oxide by converting NO
to peroxynitrite. The resulting depletion of BH4 by reaction with
peroxynitrite results in the "uncoupling" of nitric oxide synthases
so that they form superoxide rather than NO. This adds to the
oversupply of superoxide and prolongs the depletion of NO. Addition
of exogenous BH4 can reverse this uncoupling phenomenon, restoring
the production of NO and reducing the level of oxidative stress in
tissues. This mechanism is expected to complement the actions of
compounds of the invention, which reduce oxidative stress by other
means, as discussed above and throughout this invention.
VIII. EXAMPLES
The following examples are included to demonstrate preferred
embodiments of the invention. It should be appreciated by those of
skill in the art that the techniques disclosed in the examples
which follow represent techniques discovered by the inventor to
function well in the practice of the invention, and thus can be
considered to constitute preferred modes for its practice. However,
those of skill in the art should, in light of the present
disclosure, appreciate that many changes can be made in the
specific embodiments which are disclosed and still obtain a like or
similar result without departing from the spirit and scope of the
invention.
Example 1
Methods and Materials
Nitric Oxide Production and Cell Viability.
RAW264.7 macrophages were pre-treated with DMSO or drugs for 2
hours, then treated with recombinant mouse IFN.gamma. (Sigma) for
24 hours. NO concentration in media was determined using the Griess
reagent system (Promega). Cell viability was determined using WST-1
reagent (Roche).
STAT3 Phosphorylation.
HeLa cells were treated with the indicated compounds and
concentrations for 6 hours and subsequently stimulated with 20
ng/ml recombinant human IL-6 (R&D Systems) for 15 minutes.
Lysates were immunoblotted with antibodies against phosphorylated
or total STAT3 (Cell Signaling).
NF-.kappa.B Activation.
HeLa cells were transfected with pNF-.kappa.B-Luc (inducible,
Stratagene) and pRL-TK (constitutive, Promega) reporter plasmids.
Twenty-four hours later cells were pre-treated with the indicated
compounds for 2 hours. DMSO served as a vehicle control. Following
pre-treatment, cells were stimulated with 20 ng/ml recombinant
human TNF.alpha. (BD Biosciences) for 3 hours. Reporter activity
was measured using DualGlo luciferase reporter system (Promega) and
pNF-.kappa.B luciferase activity was normalized against pRL-TK
luciferase activity. Fold-induction of mean luciferase activity
relative to unstimulated (-TNF.alpha.) samples is shown. Error bars
represent the SD of the mean of 6 samples.
I.kappa.B.alpha. Degradation.
HeLa cells were treated with indicated compounds and concentrations
for 6 hours and subsequently stimulated with 20 ng/ml TNF.alpha.
for 15 minutes. Lysates were blotted with antibodies against
I.kappa.B.alpha. (Santa Cruz) and actin (Chemicon).
COX-2 Induction Western Blot.
RAW264.7 cells were pre-treated for 2 hours with indicated
compounds and subsequently stimulated with 10 ng/ml IFN.gamma. for
an additional 24 hours. COX-2 protein levels were assayed by
immunoblotting using an antibody from Santa Cruz. Actin was used as
a loading control.
Nrf2 Target Gene Induction.
MDA-MB-435 human melanoma cells were treated with vehicle (DMSO) or
the indicated compounds and concentrations for 16 hours. HO-1,
thioredoxin reductase-1 (TrxR1), .gamma.-glutamylcysteine
synthetase (.gamma.-GCS), and ferritin heavy chain mRNA levels were
quantified using qPCR and were normalized relative to a
DMSO-treated sample run in parallel. Values are averages of
duplicate wells. Primer sequences are as follows.
TABLE-US-00004 (SEQ ID NO: 1) HO-1 FW: TCCGATGGGTCCTTACACTC, (SEQ
ID NO: 2) HO-1 REV: TAGGCTCCTTCCTCCTTTCC, (SEQ ID NO: 3) TrxR1 FW:
GCAGCACTGAGTGGTCAAAA, (SEQ ID NO: 4) TrxR1 REV:
GGTCAACTGCCTCAATTGCT, (SEQ ID NO: 5) .gamma.-GCS FW:
GCTGTGGCTACTGCGGTATT, (SEQ ID NO: 6) .gamma.-GCS REV
ATCTGCCTCAATGACACCAT, (SEQ ID NO: 7) Ferritin HC FW:
ATGAGCAGGTGAAAGCCATC, (SEQ ID NO: 8) Ferritin HC REV:
TAAAGGAAACCCCAACATGC, (SEQ ID NO: 9) S9 FW: GATTACATCCTGGGCCTGAA,
(SEQ ID NO: 10) S9 REV: GAGCGCAGAGAGAAGTCGAT.
Comparison Compounds.
Some of the experimental results presented below and throughout
this application present data for not only the compounds discussed
above, but also for one or more of the triterpenoid derivatives
shown in the table below.
TABLE-US-00005 ##STR00039## 401 (RTA 401) ##STR00040## 402 (RTA
402) ##STR00041## 402-19 ##STR00042## 402-52 ##STR00043## 402-53
##STR00044## 402-54 ##STR00045## 402-55 ##STR00046## 402-56
##STR00047## 404 (RTA 404) ##STR00048## 63112 ##STR00049## 63324
##STR00050## 63166
Several of the above compounds, including 401, 402, 402-56 and 404
can be prepared according to the methods taught by Honda et al.
(1998), Honda et al. (2000b), Honda et al. (2002), Yates et al.
(2007), U.S. Pat. No. 6,974,801, and U.S. Provisional Applications
61/046,342, 61/046,352, 61/046,366, 61/111,269, and 61/111,294,
which are all incorporated herein by reference. The synthesis of
the other compounds may be prepared according to the methods
disclosed in one or more of the following applications filed
concurrently herewith, each of which is incorporated herein by
reference in their entireties: U.S. patent application by Eric
Anderson, Xin Jiang and Melean Visnick, entitled "Antioxidant
Inflammation Modulators: Oleanolic Acid Derivatives with Amino and
Other Modifications At C-17," filed Apr. 20, 2009; U.S. patent
application by Xin Jiang, Jack Greiner, Lester Maravetz, Stephen S.
Szucs, Melean Visnick, entitled "Antioxidant Inflammation
Modulators: Novel Derivatives of Oleanolic Acid," filed Apr. 20,
2009; U.S. patent application by Xin Jiang, Xiaofeng Liu, Jack
Greiner, Stephen S. Szucs, Melean Visnick entitled, "Antioxidant
Inflammation Modulators: C-17 Homologated Oleanolic Acid
Derivatives," filed Apr. 20, 2009.
Aqueous Solubility Determination.
The following procedure was used to obtain the aqueous solubility
results summarized in Example 8. Step 1. Determination of optimal
UV/vis wavelengths and generation of standard curves for a compound
of interest: (1) For eight standard calibration curves (one plate),
prepare 34 mL of 50:50 (v:v) universal buffer:acetonitrile in a 50
mL tube. (2) Using a multichannel pipet, dispense (in .mu.L) the
buffer:acetonitrile in a deep well plate as follows:
TABLE-US-00006 1 2 3 4 5 6 7 8 9 10 11 12 A 285 285 380 380 285 285
285 285 285 285 285 285 B C D E F G H
(3) Using a multichannel pipet, dispense DMSO into the same plate
as follows:
TABLE-US-00007 1 2 3 4 5 6 7 8 9 10 11 12 A 12 .mu.L 12 .mu.L 15
.mu.L 15 .mu.L 15 .mu.L 15 .mu.L 15 .mu.L 15 .mu.L 15 .mu.L 15
.mu.L B C D E F G H
(4) Add 10 mM compound in DMSO into the plates as follows:
TABLE-US-00008 1 2 3 4 5 6 7 8 9 10 11 12 A 15 .mu.L 15 .mu.L 8
.mu.L 8 .mu.L cmpd1 cmpd1 cmpd1 cmpd1 B 15 .mu.L 15 .mu.L 8 .mu.L 8
.mu.L cmpd2 cmpd2 cmpd2 cmpd2 C 15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L
cmpd3 cmpd3 cmpd3 cmpd3 D 15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L cmpd4
cmpd4 cmpd4 cmpd4 E 15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L cmpd5 cmpd5
cmpd5 cmpd5 F 15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L cmpd6 cmpd6 cmpd6
cmpd6 G 15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L cmpd7 cmpd7 cmpd7 cmpd7 H
15 .mu.L 15 .mu.L 8 .mu.L 8 .mu.L cmpd8 cmpd8 cmpd8 cmpd8
(5) Mix columns 1 and 2 by pipetting each up and down 10 times. Mix
columns 3 and 4 by pipetting up and down 10 times. Serially dilute
as follows (pipet up and down 10 times after each transfer):
TABLE-US-00009 ##STR00051## Note columns 11 and 12 contain DMSO
only and so compound should not be transferred to these wells.
(6) Cover plate with lid and shake (200-300 rpm) at room
temperature for 20 minutes. (7) Mix all wells by pipetting up and
down 10 times. (8) Transfer 120 .mu.L from each well to a UV
transparent plate. Cover and shake for 3-5 minutes. Remove any
bubbles in the wells using a pipet. (9) Read from 220 nm to 500 nm
at 10 nm increments on a spectrophotometer (e.g.,
SpectraMax.RTM.).
Step 2. Compound Solubility Testing Procedures using the
Millipore.TM. Multiscreen.RTM. Solubility Filter Plate.
Consumables: Millipore.TM. Multiscreen.RTM. Solubility Filter Plate
#MSSLBPC10 Greiner.RTM. 96 well disposable UV-Star analysis plate,
VWR#655801 Greiner.RTM. 96 well polypropylene V-bottom collection
plate, VWR#651201
Universal Aqueous Buffer: (a) To prepare 500 mL of universal
buffer, add the following: 250 mL Nanopure water; 1.36 mL (45 mM)
ethanolamine; 3.08 g (45 mM) potassium dihydrogen phosphate; 2.21 g
(45 mM) potassium acetate; thoroughly mix. (b) Adjust pH to 7.4
with HCl and q.s. to 500 mL with 0.15 M KCl. (c) Filter to remove
particulates and reduce bacterial growth. (d) Store at 4.degree. C.
in the dark.
Solubility Protocol: (a) Add 285 .mu.L of Universal Aqueous Buffer
to desired wells of the Millipore.TM. Multiscreen.RTM. Solubility
filter plate. (b) Add 15 .mu.L of 10 mM compound in DMSO to the
appropriate wells. Add 15 .mu.L of 100% DMSO only to 6 wells of the
filter plate for blanks (c) Using a multichannel pipet, mix wells
by pipetting up and down 10 times. Be careful not to touch the
filters in the plate with the tips. (d) Cover and gently shake
(200-300 rpm) filter plate for 90 minutes at room temperature. (e)
Vacuum filter the aqueous solution from the Multiscreen.RTM.
solubility filter plate into a polypropylene V-bottom plate. (f)
Transfer 60 .mu.l of filtrate to a UV transparent plate
(Greiner.RTM. UV-Star Analysis Plate). (g) Add 60 .mu.L of
acetonitrile to each well and mix by pipetting up and down 10
times. (h) Cover and gently shake for 3-5 minutes. Remove any
bubbles with a pipet. (i) Measure the absorbance of each well in
the plate on the spectrophotometer (UV/vis) at the desired
wavelength. For compounds in a plate with different absorbance
peaks, set the spectrophotometer to read a spectrum (e.g., from 220
nm to 460 nm). (j) Identify concentration using measured absorbance
for each compound and the predetermined standard curve (see Step
1).
Example 2
Synthesis of Oleanolic Acid Derivatives
The synthesis of compounds 402-02 and 402-51 started from compound
1 (Scheme 1). Compound 1 was oxidized with bleach to give ketone 2
in 80% yield. Formylation of 2 with ethyl formate using sodium
methoxide as the base afforded compound 402-48 (70% yield), which
was then treated with hydroxylamine hydrochloride in aqueous EtOH
at 55.degree. C. to give isoxazole 402-49 in 93% yield. Cleavage of
the isoxazole under basic conditions gave .alpha.-cyanoketone
402-46 in quantitative yield as a mixture of ketone and enol forms.
Compound 402-46 was treated with 1,3-dibromo-5,5-dimethylhydantoin,
followed by elimination of HBr using pyridine as the base, to give
compound 402-02 in 81% yield (from 402-49), which was demethylated
with LiI in refluxing DMF to give acid 402-51 in 95% yield.
##STR00052## ##STR00053##
Compound 402-63 was oxidized with Dess-Martin periodinane to give
aldehyde 402-64 in 47% yield (Scheme 2).
##STR00054##
The synthesis of compounds 402-59 and 402-57 began from 402-51.
Compound 402-51 was treated with oxalyl chloride and catalytic DMF
to give acid chloride 3. Compound 3 was treated with ammonia in
methanol to give 402-59 (99% from 402-51). Dehydration of 402-59
utilizing TFAA and Et.sub.3N gave dicyano compound 402-57 (45%
yield).
##STR00055##
404-02 was synthesized from compound 3 as summarized in Scheme 4.
Compound 3 was reacted with 2,2,2-trifluoroethylamine-HCl in
toluene and water at 70.degree. C. with NaHCO.sub.3 as the base,
giving 404-02 in 69% yield.
##STR00056##
##STR00057##
The synthesis of 402-66 began from isoxazole compound 402-49.
Reduction of the ketone in 402-49 was achieved by treatment with
LiAlH.sub.4 in THF at 0.degree. C., giving compounds 4 and 5 (as a
1:1 mixture of diastereotopic alcohols). Compound 4 was treated
with NaOMe in MeOH at 55.degree. C. to give 6, which exists as a
3:2 mixture of keto and enol tautomeric forms. Bromination and
subsequent dehydrobromination of 6 by treatment with
1,3-dibromo-5,5-dimethylhydantoin and then pyridine gave 402-66 in
33% yield (from 4). Using the same synthetic sequence, compound 5
was converted into 63219 in 28% overall yield.
##STR00058## ##STR00059##
##STR00060## ##STR00061##
##STR00062##
##STR00063##
##STR00064##
##STR00065## ##STR00066## ##STR00067##
##STR00068##
##STR00069##
##STR00070##
##STR00071##
##STR00072##
##STR00073##
##STR00074##
##STR00075##
Example 3
Synthesis and Characterization of Oleanolic Acid Derivatives
Compound 2:
Bleach (5.25 wt % of NaClO (aq), 129 mL, 91 mmol) was added to a
stirred solution of compound 1 (34.67 g, 71 mmol) in AcOH (471 mL)
at room temperature. After stirring for 40 min, the reaction
mixture was poured into ice-water (1.5 L) and stirred for 5 min.
The white precipitate was collected by filtration and washed
thoroughly with water. The filtered solid was then dissolved in
EtOAc and washed with NaHCO.sub.3 (aq) solution, dried with
MgSO.sub.4 and concentrated. The residue obtained was purified by
column chromatography (silica gel, 10% to 25% EtOAc in hexanes) to
give product 2 (27.8 g, 80%) as a white foam solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 3.69 (s, 3H), 2.80 (m, 1H), 2.65 (d,
1H, J=4.0 Hz), 2.53 (ddd, 1H, J=7.2, 10.8, 16.0 Hz), 2.38 (ddd, 1H,
J=3.6, 6.8, 16.0 Hz), 2.16-2.30 (m, 2H), 1.95 (m, 1H), 1.89 (m,
1H), 1.80 (m, 2H), 1.62-1.73 (m, 3H), 1.57 (m, 2H), 1.47 (m, 2H),
1.15-1.40 (m, 7H), 1.09 (s, 3H), 1.05 (s, 3H), 1.01 (s, 3H), 0.99
(s, 3H), 0.98 (s, 3H), 0.95 (s, 3H), 0.90 (s, 3H); m/z 485.3
(M+1).
Compound 402-48:
NaOMe solution (25% w/w in MeOH, 132.3 mL, 570 mmol) was added to a
solution of compound 2 (27.6 g, 57 mmol) in MeOH (250 mL) under
nitrogen. The reaction mixture was heated to 55.degree. C. in an
oil bath, and HCO.sub.2Et (93 mL, 1.15 mmol, 20 eq) was added
dropwise via an addition funnel. The reaction mixture was stirred
at 55.degree. C. for 24 h and then at room temperature for another
40 h. After removal of MeOH (150 mL) by evaporation, t-BuOMe (200
mL) was added, and the mixture was cooled to 0.degree. C. 12 N HCl
(aq) (50 mL, 600 mmol, 10.5 eq) was then added over 10 min, and the
mixture was extracted with EtOAc. The combined extracts were washed
with water, dried with MgSO.sub.4, and concentrated. The brown oil
obtained was purified by column chromatography (silica gel, 5% to
10% EtOAc in hexanes) to give product 402-48 (20.5 g, 70%) as a
white foam solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 14.89
(d, 1H, J=3.2 Hz), 8.61 (d, 1H, J=3.2 Hz), 3.69 (s, 3H), 2.80 (m,
1H), 2.67 (d, 1H, J=4.0 Hz), 2.20-2.34 (m, 3H), 1.98 (m, 1H),
1.62-1.92 (m, 6H), 1.10-1.56 (m, 10H), 1.20 (s, 3H), 1.12 (s, 3H),
1.02 (s, 3H), 0.99 (s, 3H), 0.96 (s, 3H), 0.91 (s, 3H), 0.85 (s,
3H); m/z 513.3 (M+1).
Compound 402-49:
A mixture of compound 402-48 (20.3 g, 40 mmol) and NH.sub.2OH.HCl
(4.12 g, 59 mmol) in EtOH (300 mL) and water (60 mL) was heated at
55.degree. C. for 14 h. After cooling to room temperature, EtOH was
removed by evaporation, and the white slurry obtained was extracted
with EtOAc. The combined extracts were washed with water, dried
with MgSO.sub.4, and concentrated. The residue obtained was
purified by column chromatography (silica gel, 10% to 20% EtOAc in
hexanes) to give product 402-49 (18.8 g, 93%) as a white solid:
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.99 (s, 1H), 3.70 (s,
3H), 2.81 (m, 1H), 2.68 (d, 1H, J=4.4 Hz), 2.37 (d, 1H, J=15.2 Hz),
2.23-2.33 (m, 2H), 1.76-1.98 (m, 5H), 1.68 (m, 3H), 1.11-1.62 (m,
9H), 1.32 (s, 3H), 1.23 (s, 3H), 1.02 (s, 3H), 0.99 (s, 3H), 0.97
(s, 3H), 0.91 (s, 3H), 0.84 (s, 3H); m/z 510.3 (M+1).
Compound 402-46:
NaOMe (25% w/w in MeOH, 8.75 mL, 38 mmol) was added dropwise to a
suspension of 402-49 (16.16 g, 31.7 mmol) in MeOH (55 mL) at
0.degree. C. under N.sub.2. The reaction mixture was heated at
55.degree. C. for 2 h and then cooled to 0.degree. C. t-BuOMe (150
mL) and 1 N HCl (aq) (50 mL) were added successively, and the
mixture was extracted with EtOAc. The combined extracts were washed
with water, dried with MgSO.sub.4, and concentrated to give
compound 402-46 (17.80 g, 100%) as a white foam solid. 402-46 is a
mixture of two equilibrium forms, the enol form (as shown in Scheme
1) and the ketone form, in the ratio of 2:3. .sup.1H NMR of the
mixture: (400 MHz, CDCl.sub.3) .delta. 5.69 (s, 0.4H), 3.87 (m,
0.6H), 2.80 (m, 1H), 2.65 (m, 1H), 0.82-2.30 (m, 44H); m/z 510.3
(M+1).
Compound 402-02:
1,3-Dibromo-5,5-dimethylhydantoin (5.98 g, 20.9 mmol) was added to
a solution of compound 402-46 (17.76 g, 35 mmol) in DMF (75 mL) at
10.degree. C. After stirring at room temperature for 2 h, pyridine
(8.5 mL, 105 mmol) was added, and the reaction mixture was heated
at 55.degree. C. for 15 h. After cooling to room temperature, the
mixture was poured into water (700 mL) and stirred for 5 min. The
pale brown precipitate was collected by filtration and washed with
water. The solid was dissolved in CH.sub.2Cl.sub.2, and the
solution was washed with 1 N HCl (aq) and water, then dried with
MgSO.sub.4 and concentrated. The residue obtained was purified by
column chromatography (silica gel, 0% to 70% EtOAc in hexanes) to
give product 402-02 (14.3 g, 81%) as a white solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 7.65 (s, 1H), 3.69 (s, 3H), 2.82 (m,
1H), 2.68 (d, 1H, J=4.4 Hz), 2.44 (dd, 1H, J=4.8, 16.0 Hz), 2.35
(dd, 1H, J=12.8, 16.0 Hz), 1.86-2.00 (m, 3H), 1.81 (m, 1H),
1.60-1.71 (m, 4H), 1.42-1.55 (m, 3H), 1.24 (m, 1H), 1.10-1.24 (m,
4H), 1.22 (s, 3H), 1.16 (s, 3H), 1.15 (s, 3H), 1.07 (s, 3H), 0.99
(s, 3H), 0.97 (s, 3H), 0.92 (s, 3H); m/z 508.2 (M+1).
Compound 402-51:
A stream of nitrogen was bubbled through a stirring solution of
compound 402-02 (6.31 g, 12.4 mmol) and LiI (33.35 g, 248 mmol) in
DMF (87 mL) at 160.degree. C. for 8 h. After cooling to 50.degree.
C., the reaction mixture was diluted with EtOAc (100 mL). 1 N HCl
(aq) solution (30 mL) was then added at room temperature and
stirred for 5 min. The mixture was extracted with EtOAc, and the
combined extracts were washed with water, 10%
Na.sub.2S.sub.2O.sub.3 (aq) and water, then dried with MgSO.sub.4
and concentrated. The residue obtained was purified by column
chromatography (silica gel, 5% to 50% EtOAc in CH.sub.2Cl.sub.2) to
give acid 402-51 (6.02 g, 95%) as a white solid: .sup.1H NMR (400
MHz, CDCl.sub.3) .delta. 10.41 (bs, 1H), 7.65 (s, 1H), 2.80 (m,
1H), 2.74 (d, 1H, J=4.4 Hz), 2.46 (dd, 1H, J=4.8, 16.0 Hz), 2.37
(dd, 1H, J=12.8, 16.0 Hz), 1.86-2.02 (m, 4H), 1.44-1.79 (m, 8H),
1.35 (m, 1H), 1.12-1.29 (m, 3H), 1.22 (s, 3H), 1.16 (s, 3H), 1.14
(s, 3H), 1.11 (s, 3H), 0.99 (s, 3H), 0.98 (s, 3H), 0.93 (s, 3H);
m/z 494.3 (M+1).
Compound 402-64:
NaHCO.sub.3 (78 mg, 0.93 mmol) and Dess-Martin periodinane (99 mg,
0.23 mmol) were added successively to a solution of 402-63 (45 mg,
94 .mu.mol) in CH.sub.2Cl.sub.2 (5 mL) at room temperature. After
stirring for 1 h, 5% Na.sub.2S.sub.2O.sub.3 (aq) solution was
added. The reaction mixture was extracted with t-BuOMe, and the
combined extracts were washed with NaHCO.sub.3 (aq) solution, dried
with MgSO.sub.4, and concentrated. The crude product obtained was
purified by column chromatography (silica gel, 0% to 35% EtOAc in
hexanes) to give 402-64 (22 mg, 49%) as a white foam solid: .sup.1H
NMR (300 MHz, CDCl.sub.3) .delta. 9.33 (s, 1H), 7.62 (s, 1H), 2.61
(m, 1H), 2.50 (d, 1H, J=4.4 Hz), 2.45 (dd, 1H, J=4.8, 16.4 Hz),
2.34 (dd, 1H, J=13.2, 16.4 Hz), 1.92-2.00 (m, 2H), 1.88 (m, 1H),
1.42-1.74 (m, 9H), 1.28-1.35 (m, 2H), 1.21 (s, 3H), 1.20 (m, 1H),
1.15 (s, 3H), 1.14 (s, 3H), 1.12 (m, 1H), 1.06 (s, 3H), 0.97 (s,
6H), 0.93 (s, 3H); m/z 478.2 (M+1).
Compound 402-59:
To a solution of 402-51 (2.08 g, 4.21 mmol) in CH.sub.2Cl.sub.2 (28
mL) were successively added oxalyl chloride (1.07 mL, 12.64 mmol)
and DMF (5 drops, cat.) at 0.degree. C. The reaction was allowed to
warm to room temperature and was stirred for 3 h. The reaction
mixture was concentrated and dried in vacuo 30 min, to give acid
chloride 3 as a yellow solid, which was used directly in the next
step. To a solution of 3 (2.16 g, 4.21 mmol) in THF (28 mL) at
0.degree. C. was added ammonia (2.0 M solution in MeOH, 11 mL,
22.00 mmol). The reaction was allowed to warm to room temperature
and was stirred for 5 h. The solvents were then evaporated, and the
residue was extracted with EtOAc. The extracts were washed with
water, 1 N HCl (aq), and water, then dried over MgSO.sub.4,
filtered, and concentrated to give 402-59 (2.06 g, 99%) as a pale
yellow solid. A small amount (53 mg) was purified by column
chromatography (silica gel, 0% to 25% EtOAc in CH.sub.2Cl.sub.2) to
give higher purity 402-59 (14 mg, white solid) for biological
assay: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.65 (s, 1H), 5.63
(br s, 1H), 5.36 (br s, 1H), 2.90 (br d, 1H, J=5.2 Hz), 2.71 (br d,
1H, J=12 Hz), 2.42 (m, 2H), 1.96-2.10 (m, 4H), 1.78-1.90 (m, 2H),
1.45-1.69 (m, 6H), 1.23-1.40 (m, 4H), 1.22 (s, 3H), 1.16 (s, 3H),
1.15 (s, 3H), 1.13 (s, 3H), 0.99 (s, 3H), 0.98 (s, 3H), 0.93 (s,
3H); m/z 493.3 (M+1)
Compound 402-57:
A solution of 402-59 (2.01 g, 4.08 mmol) in CH.sub.2Cl.sub.2 (28
mL) was prepared and cooled to 0.degree. C. To this solution were
added TFAA (0.91 mL, 6.55 mmol) and Et.sub.3N (1.48 mL, 10.62
mmol). The reaction was stirred at 0.degree. C. for 3 h, after
which it was quenched by the addition of saturated NaHCO.sub.3 (aq)
solution (40 mL). After stirring for 10 min, the reaction mixture
was extracted with CH.sub.2Cl.sub.2 and washed with saturated
NaHCO.sub.3 (aq), water, 1 N HCl (aq), and water. The extracts were
dried over MgSO.sub.4, filtered, and concentrated. The crude
product was purified by column chromatography (silica gel, 5% to
35% EtOAc in hexanes). The purified product was triturated with
EtOH, then filtered and dried on the filter to give 402-57 (0.87 g,
45%) as a powdery white solid: .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 7.63 (s, 1H), 3.04 (d, 1H, J=4.4 Hz), 2.38-2.57 (m, 3H),
1.91-2.19 (m, 5H), 1.61-1.78 (m, 4H), 1.44-1.54 (m, 3H), 1.32 (s,
3H), 1.26-1.30 (m, 4H), 1.22 (s, 3H), 1.19 (s, 3H), 1.16 (s, 3H),
0.99 (s, 3H), 0.95 (s, 3H), 0.92 (s, 3H); m/z 475.2 (M+1).
Compound 404-02:
To a solution of 3 (3.03 g, 5.92 mmol) in toluene (84 mL) was added
NaHCO.sub.3 (1.98 g). A solution of trifluoroethylamine
hydrochloride (5.64 g, 41.62 mmol) in water (14 mL) was prepared,
then added to the reaction. The reaction was heated to 70.degree.
C. and stirred for 2 h. After cooling to room temperature, the
reaction mixture was extracted with EtOAc and washed with brine.
The combined extracts were dried over MgSO.sub.4, filtered, and
concentrated. The crude product was purified by column
chromatography (silica gel, 0% to 40% EtOAc in CH.sub.2Cl.sub.2) to
give 404-02 (2.35 g, 69%) as a white solid: .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.65 (s, 1H), 5.94 (br t, 1H, J=8 Hz), 4.10 (m,
1H), 3.69-3.88 (m, 1H), 2.84 (d, 1H, J=8 Hz), 2.78 (br d, 1H, J=16
Hz), 2.38 (m, 2H), 2.12 (m, 1H), 2.06 (m, 2H), 1.61-1.83 (m, 5H),
1.24-1.52 (m, 8H), 1.22 (s, 3H), 1.16 (s, 3H), 1.14 (s, 3H), 1.07
(s, 3H), 0.99 (s, 6H), 0.93 (s, 3H); m/z 575.3 (M+1).
Compounds 4, 5:
To a solution of 402-49 (395 mg, 0.775 mmol) in THF (7.8 mL) was
added LiAlH.sub.4 (1.0 M solution in THF, 0.78 mL, 0.780 mmol) at
0.degree. C. The reaction was stirred at 0.degree. C. for 40 min,
after which it was quenched by the addition of water (5 mL) and
stirred 5 min. The reaction mixture was extracted with EtOAc and
washed with water. Solid NaCl was added to break up emulsions. The
combined extracts were dried over Na.sub.2SO.sub.4, filtered, and
concentrated. The crude product was purified by column
chromatography (silica gel, 10% to 70% EtOAc in hexanes) to give
both 4 (151 mg, 38%) as a white solid and to give 5 (134 mg, 34%)
as a white solid:
Compound 4: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.99 (s, 1H),
3.79 (m, 1H), 3.72 (s, 3H), 2.75 (m, 1H), 2.52 (d, 1H, J=14.4 Hz),
1.95-2.11 (m, 2H), 1.57-1.88 (m, yyH), 1.24-1.54 (m, yyH), 1.30 (s,
3H), 1.20 (s, 3H), 1.00 (s, 3H), 0.93 (s, 6H), 0.92 (s, 3H), 0.82
(s, 3H); m/z 512.3 (M+1).
Compound 5: m/z 494.3 (M-17), 434.3 (M-17-60).
Compound 6:
To a solution of 4 (371 mg, 66 .mu.mol) in MeOH (7.3 mL) was added
NaOMe (25 wt % solution in MeOH, 0.42 mL, 1.837 mmol) at room
temperature. The reaction was heated to 55.degree. C. and stirred 7
h. After cooling the reaction to room temperature, the reaction
mixture was diluted with MTBE (10 mL), then quenched with 1 N HCl
(aq) (10 mL). The reaction mixture was extracted with EtOAc and
washed with 1 N HCl (aq) and brine. The combined extracts were
dried over Na.sub.2SO.sub.4, filtered, and concentrated to give 6
(361 mg, 94%) as a white solid. Compound 6 is a mixture of two
equilibrium forms, the enol form (as shown in Scheme 5) and the
ketone form, in the ratio of 2:3. .sup.1H NMR of the mixture: (400
MHz, CDCl.sub.3) .delta. 5.66 (d, 0.4H, J=4.8 Hz), 4.09 (br, 1H),
3.90 (m, 0.6H), 3.71 (s, 1.2H), 3.68 (s, 1.8H), 2.73 (m, 1H), 2.48
(m, 1H), 2.14-2.26 (m, 2H), 0.80-2.02 (m, 39H); m/z 494.3 (M-17),
434.3 (M-77).
Compound 402-66:
A solution of 6 (361 mg, 0.705 mmol) in DMF (7.1 mL) was prepared.
1,3-Dibromo-5,5-dimethylhydantoin (120 mg, 0.420 mmol) was added,
and the reaction was stirred at room temperature for 1 h. Pyridine
(0.23 mL, 2.858 mmol) was added, and the reaction was heated to
55.degree. C. and stirred 10 h. After cooling the reaction to room
temperature, the reaction mixture was extracted with EtOAc and
washed with 5% Na.sub.2S.sub.2O.sub.3 (aq), water, 1 N HCl (aq),
and water. The EtOAc extracts were dried over Na.sub.2SO.sub.4,
filtered, and evaporated. The crude product was purified by column
chromatography (silica gel, 5% to 40% EtOAc in hexanes) to give
402-66 (127 mg, 35% yield) as a white solid; .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.81 (s, 1H), 3.81 (ddd, 1H, J=4.8, 10.8, 15.6
Hz), 3.72 (s, 3H), 2.75 (m, 1H), 2.01 (m, 2H), 1.74-1.88 (m, 3H),
1.24-1.72 (m, 15H), 1.19 (s, 3H), 1.13 (s, 3H), 1.12 (s, 3H), 0.98
(s, 3H), 0.96 (s, 3H), 0.95 (s, 3H), 0.94 (s, 3H); m/z 492.3
(M-17), 432.3 (M-77).
Compound 7:
Using the procedure described for the synthesis of compound 6 from
compound 4, compound 7 (96 mg, 89% yield) was produced from
compound 5 (108 mg, 0.211 mmol): m/z 494.3 (M-17).
Compound 63219:
Using the procedure described for the synthesis of compound 402-66
from compound 6, compound 63219 (30 mg, 32% yield) was produced
from compound 7 (95 mg, 0.186 mmol): .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.81 (1H, s), 4.16 (1H, bs), 3.68 (3H, s),
2.44-2.54 (2H, m), 1.98-2.10 (2H, m), 1.78-1.94 (4H, m), 1.42-1.76
(7H, m), 1.00-1.42 (6H, m), 1.32 (3H, s), 1.23 (3H, s), 1.13 (3H,
s), 1.11 (3H, s), 0.94 (3H, s), 0.93 (3H, s), 0.92 (3H, s); m/z
492.3 (M-17).
Compound 8:
Oxalyl chloride (0.11 mL, 1.30 mmol) and catalytic amount of DMF
were added sequentially to a solution of compound 402-51 (200 mg,
0.41 mmol) in CH.sub.2Cl.sub.2 (4 mL) at 0.degree. C. The reaction
mixture was warmed to room temperature and stirred for 2 h. After
removing the solvent by evaporation, the crude acid chloride was
obtained as a light yellow foam solid. Hydrazine hydrate (64% of
hydrazine, 0.50 mL) was added to a solution of acid chloride in
Et.sub.2O (8 mL) at 0.degree. C. After stirring for 30 min,
CH.sub.2Cl.sub.2 was added. The mixture was washed with water,
dried over MgSO.sub.4, filtered and evaporated to give compound 8
(200 mg, 97% yield) as white solid, which was used in the next step
without further purification: m/z 508.3 (M+1).
Compound 9:
Et.sub.3N (0.12 mL, 0.86 mmol) and acetyl chloride (37 .mu.L, 0.52
mmol) were added sequentially to a solution of compound 8 (200 mg,
0.39 mmol) in CH.sub.2Cl.sub.2 (4 mL) at r.t. After stirring for 30
min, Et.sub.3N (0.36 mL, 2.59 mmol) and acetyl chloride (110 .mu.L,
1.55 mmol) were added again. After stirring for another 30 min,
NaHCO.sub.3 (aq.) solution was added to quench the reaction. The
reaction mixture was transferred to a separatory funnel, and
extracted with EtOAc. The combined extracts were washed with water,
dried over MgSO.sub.4, filtered and evaporated. The residue was
purified by silica gel chromatography (0% to 75% EtOAc in hexanes)
to give compound 9 (180 mg, 77% yield) as a white foam solid: m/z
592.3 (M+1).
Compound 63264:
NaOMe (25% w/w in MeOH, 0.14 mL, 0.61 mmol) was added to a solution
of compound 9 (180 mg, 0.30 mmol) in MeOH (3 mL) at 0.degree. C.
After stirring at r.t. for 10 min, the reaction mixture was treated
with t-BuOMe (10 mL) and 1N HCl (aq.) (1 mL), which was then
transferred to a separatory funnel and extracted with EtOAc. The
combined extracts were washed with NaHCO.sub.3 (aq.) solution,
dried over MgSO.sub.4, filtered and evaporated. The residue was
purified by silica gel chromatography (0% to 100% EtOAc in hexanes)
to give compound 63264 (121 mg, 72% yield) as a white solid:
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.98 (d, 1H, J=4.4 Hz),
7.77 (d, 1H, J=4.4 Hz), 7.65 (s, 1H), 2.89 (d, 1H, J=4.4 Hz), 2.82
(m, 1H), 2.34-2.45 (m, 2H), 2.10 (m, 1H), 2.08 (s, 3H), 1.82-2.02
(m, 4H), 1.60-1.69 (m, 3H), 1.44-1.53 (m, 4H), 1.16-1.40 (m, 4H),
1.22 (s, 3H), 1.16 (s, 3H), 1.15 (s, 3H), 1.13 (s, 3H), 0.99 (s,
3H), 0.98 (s, 3H), 0.94 (s, 3H); m/z 550.3 (M+1).
Compound 63267:
A solution of compound 63264 (74 mg, 0.13 mmol), TsOH (13 mg, 0.068
mmol) in toluene (5 mL) was heated at reflux with dean-stark trap
for 1 h. After cooling to r.t., the reaction mixture was
transferred to a separatory funnel, washed with NaHCO.sub.3 (aq.)
solution, dried over MgSO.sub.4, filtered and evaporated. The
residue was purified by silica gel chromatography (0% to 100% EtOAc
in hexanes) to give compound 63267 (24 mg, 33% yield) as a white
foam solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.64 (s, 1H),
2.94 (m, 1H), 2.79 (d, 1H, J=4.4 Hz), 2.54 (s, 3H), 2.46 (m, 1H),
2.34 (m, 1H), 2.21 (m, 1H), 1.84-2.06 (m, 5H), 1.56-1.70 (m, 4H),
1.24-1.47 (m, 5H), 1.23 (s, 3H), 1.18 (m, 1H), 1.15 (s, 3H), 1.13
(s, 3H), 1.05 (s, 3H), 1.02 (s, 3H), 0.98 (s, 3H), 0.93 (s, 3H);
m/z 532.3 (M+1).
Compound 11:
Et.sub.3N (1.46 mL, 10.49 mmol) and TFAA (0.88 mL, 6.33 mmol) were
added sequentially to a solution of compound 10 (1.97 g, 4.20 mmol)
in CH.sub.2Cl.sub.2 (42 mL) at 0.degree. C. After stirring for 1.5
h, NaHCO.sub.3 (aq.) solution was added to the reaction mixture,
which was then transferred to a separatory funnel and extracted
with CH.sub.2Cl.sub.2. The combined extracts were dried over
MgSO.sub.4, filtered and evaporated. The residue was purified by
silica gel chromatography (0% to 35% EtOAc in hexanes) to give
compound 11 (1.62 g, 85% yield) as a white solid: m/z 452.3.
Compound 12:
A solution of Bu.sub.3SnN.sub.3 (1.00 mL, 3.62 mmol) and compound
11 (1.36 g, 3.02 mmol) in xylene (5.0 mL) was heated at reflux for
48 h. After cooling to r.t., the reaction mixture was purified by
silica gel chromatography (0% to 30% EtOAc in CH.sub.2Cl.sub.2) to
give compound 12 (994 mg, 67% yield) as a light yellow foam solid:
m/z 493.3 (M+1).
Compound 13:
NaOMe solution (25% w/w in MeOH, 1.16 mL, 5.07 mmol) was added
dropwise to a mixture of compound 12 (168 mg, 0.34 mmol) and
HCO.sub.2Et (0.82 mL, 10.19 mmol) at 0.degree. C. under N.sub.2.
After stirring at room temperature for 1 h, t-BuOMe (10 mL) was
added. The mixture was cooled to 0.degree. C., and 12 N HCl (aq)
(0.42 mL, 5.04 mmol) was added slowly. The mixture was transferred
to a separatory funnel and extracted with EtOAc. The combined
extracts were washed with water, dried over MgSO.sub.4 and
concentrated to give crude 2-formyl ketone, which was then mixed
with NH.sub.2OH.HCl (36 mg, 0.51 mmol), EtOH (4 mL), and water (0.4
mL), and heated at 60.degree. C. for 3 h. After removing EtOH by
evaporation, the white slurry obtained was transferred to a
separatory funnel and extracted with CH.sub.2Cl.sub.2. The combined
extracts were washed with water, dried over MgSO.sub.4, and
concentrated. The residue was purified by column chromatography
(silica gel, 0% to 30% EtOAc in CH.sub.2Cl.sub.2) to give compound
13 (95 mg, 54% yield) as a white foam solid: m/z 520.3 (M+1).
Compound 63229:
Using the procedure described for the synthesis of compound 402-66
from compound 4, 63229 (12 mg, 60% yield) was produced from
compound 13 (20 mg, 0.038 mmol): .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 7.70 (s, 1H), 3.03 (m, 1H), 2.69 (d, 1H, J=4.0 Hz), 2.52
(dd, 1H, J=4.4, 16.8 Hz), 2.29-2.36 (m, 2H), 1.96-2.03 (m, 3H),
1.56-1.82 (m, 6H), 1.25-1.57 (m, 6H), 1.22 (s, 3H), 1.18 (m, 1H),
1.13 (s, 3H), 1.11 (s, 3H), 1.04 (s, 3H), 1.03 (s, 3H), 0.98 (s,
3H), 0.75 (s, 3H); m/z 518.3 (M+1).
Compound 14:
TMSCHN.sub.2 (2.0 M in Et.sub.2O, 89 .mu.L, 0.18 mmol) was added to
a solution of compound 13 (84 mg, 0.16 mmol) in THF (1.25 mL) and
MeOH (0.31 mL) at 0.degree. C. After stirring at room temperature
for 10 min, acetic acid was added to quench the reaction. The
reaction mixture was diluted with EtOAc, transferred to a
separatory funnel, washed with NaHCO.sub.3 (aq.) solution, dried
over MgSO.sub.4 and evaporated. The residue was purified by column
chromatography (silica gel, 0% to 60% EtOAc in hexanes) to give
compound 14 (67 mg, 77% yield) as a white solid: m/z 534.3
(M+1).
Compound 63230:
Using the procedure described for the synthesis of compound 402-66
from compound 4, 63230 (45 mg, 69% yield) was produced from
compound 14 (65 mg, 0.12 mmol): .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 7.62 (s, 1H), 4.32 (s, 3H), 3.11 (m, 1H), 2.68 (d, 1H,
J=4.4 Hz), 2.42 (dd, 1H, J=4.8, 16.4 Hz), 2.27 (dd, 1H, J=13.2,
16.4 Hz), 2.22 (dd, 1H, J=4.4, 14.8 Hz), 1.94-2.04 (m, 3H), 1.79
(m, 1H), 1.54-1.63 (m, 5H), 1.36-1.50 (m, 4H), 1.26 (m, 1H), 1.20
(s, 3H), 1.13 (m, 1H), 1.12 (s, 3H), 1.09 (s, 3H), 1.05 (s, 3H),
1.01 (s, 3H), 0.97 (s, 3H), 0.70 (s, 3H); m/z 532.3 (M+1).
Compound 63223:
Using the procedure described for the synthesis of compound 13 from
compound 12, compound 63223 (1.95 g, 47% yield) was produced from
compound 15 (3.93 g, 7.12 mmol) as a pale yellow solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 7.98 (1H, s), 5.91 (1H, t, J=6.0 Hz),
4.00-4.15 (1H, m), 3.55-3.90 (1H, m), 2.72-2.82 (2H, m), 2.20-2.40
(3H, m), 1.88-2.16 (4H, m), 1.10-1.84 (13H, m), 1.31 (3H, s), 1.21
(3H, s), 1.00 (3H, s), 0.98 (6H, s), 0.91 (3H, s), 0.82 (3H, s);
m/z 577.3 (M+H).
Compound 63227:
Using the procedure described for the synthesis of compound 6 from
compound 4, compound 63227 (1.64 g, quantitative yield) was
produced from compound 63223 (1.61 g, 2.79 mmol): .sup.1H NMR (400
MHz, CDCl3) for enol form: .delta. 5.91 (1H, t, J=6.0 Hz), 5.78
(1H, bs, enol), 4.00-4.16 (1H, m), 3.75-3.94 (1H, m), 2.70-2.85
(2H, m), 1.90-2.30 (5H, m), 0.80-1.88 (36H, m); m/z 577.3 (M+H)
(for both enol and ketone isomers).
Compound 63237:
A solution of 63227 (1.61 g, 2.79 mmol) in DMF (9.3 mL) was
prepared. 1,3-Dibromo-5,5-dimethylhydantoin (456 mg, 1.59 mmol) was
added, and the reaction was stirred at room temperature for 3 h.
Pyridine (0.67 mL, 8.33 mmol) was added, and the reaction was
heated to 55.degree. C. and stirred 16 h. After cooling the
reaction to room temperature, the reaction mixture was extracted
with EtOAc and washed with 5% Na.sub.2S.sub.2O.sub.3 (aq), 1 N HCl
(aq), and water. The EtOAc extracts were dried over
Na.sub.2SO.sub.4, filtered, and evaporated. Compound 63237 was a
minor component (18%) by crude LC-MS analysis. The crude product
was purified by column chromatography (silica gel, 5% to 35% EtOAc
in hexanes) to give 63237 (188 mg, 12%) as a yellow foam solid:
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.67 (s, 1H), 3.82 (m,
2H), 2.91 (m, 1H), 2.54 (m, 1H), 2.41 (m, 1H), 2.08 (m, 1H),
1.83-1.94 (m, 2H), 1.52-1.74 (m, 6H), 1.39 (s, 3H), 1.22 (s, 6H),
1.20-1.49 (m, 7H), 1.16 (s, 3H), 0.98 (s, 3H), 0.96 (s, 3H), 0.95
(s, 3H); m/z 573.3 (M+1).
Compound 16:
Using the procedure described for the synthesis of compound 4 from
compound 402-49, compound 16 (100 mg, 19% yield) was produced from
compound 63223 (531 mg, 0.921 mmol): m/z 579.3 (M+1)
Compound 17:
Using the procedure described for the synthesis of compound 6 from
compound 4, compound 17 (90 mg, 92% yield) was produced from
compound 16 (98 mg, 0.169 mmol): m/z 561.3 (M-17).
Compound 63268:
Using the procedure described for the synthesis of compound 402-66,
compound 63268 (50 mg, 56% yield) was produced from compound 17 (90
mg, 0.156 mmol): .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.78 (s,
1H), 6.09 (t, 1H, J=6.4 Hz), 4.06 (m, 1H), 3.89 (m, 1H), 3.78 (m,
1H), 2.70 (m, 1H), 1.98-2.05 (m, 2H), 1.84 (dd, 1H, J=4.4, 10.8
Hz), 1.75 (ddd, 1H, J=4.4, 13.6, 13.6 Hz), 1.20-1.62 (m, 15H), 1.18
(s, 3H), 1.11 (s, 3H), 1.10 (s, 3H), 1.06 (m, 1H), 0.98 (s, 3H),
0.95 (s, 3H), 0.94 (s, 3H), 0.91 (s, 3H); m/z 559.3 (M-17).
Compound 19:
NaOMe (25 w/w % solution in MeOH, 7.29 mL, 31.88 mmol) was added to
a solution of compound 18 (1.00 g, 2.12 mmol) in ethyl formate
(5.13 mL, 63.78 mmol) at 0.degree. C. After stirring for 1.5 h,
t-BuOMe (10 mL) and 12 N (aq.) HCl (2.66 mL, 31.92 mmol) were added
sequentially. After stirring for another 5 min, the reaction
mixture was transferred to a reparatory funnel, which was extracted
with EtOAc. The combined extracts were washed with water. The
organic layer was separated, which was dried over MgSO.sub.4,
filtered, and concentrated. The crude product was mixed with
NH.sub.2OH--HCl (0.22 g, 3.17 mmol), water (2 mL) and EtOH (35 mL).
The reaction mixture was heated at 65.degree. C. for 3.5 h, after
which EtOH was removed by evaporation. The residue was partitioned
between EtOAc and water. The organic extract was separated, which
was dried over MgSO.sub.4, filtered, and evaporated. The residue
was purified by silica gel chromatography (0% to 60% EtOAc in
hexanes) to give compound 19 (820 mg, 78% yield) as a white solid:
m/z 496.3 (M+1).
Compound 20:
Oxalyl chloride (110 .mu.L, 1.30 mmol) was added to a solution of
compound 19 (195 mg, 0.39 mmol) in CH.sub.2Cl.sub.2 (4 mL) at
0.degree. C., followed by the addition of catalytic amount of DMF.
The reaction was stirred at room temperature for 2 h, after which
CH.sub.2Cl.sub.2 was evaporated under vacuum to give acid chloride
as a light yellow foam solid.
Et.sub.3N (113 .mu.L, 0.81 mmol) and a solution of acethydrazide
(50 mg, 0.67 mmol) in CH.sub.2Cl.sub.2 (2 mL) were added
sequentially to a suspension of the acid chloride in ether (4 mL)
at 0.degree. C. The reaction was warmed to room temperature and
stirred for 30 min. EtOAc was then added, and the crude mixture was
transferred to a separatory funnel, which was washed with water, 1N
(aq.) HCl, water. The organic layer was separated, which was dried
over MgSO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 100% EtOAc in
hexanes) to give product 20 (215 mg, 99% yield) as a white foam
solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 8.14 (d, 1H, J=5.2
Hz), 8.05 (d, 1H, J=5.2 Hz), 8.00 (s, 1H), 2.86 (m, 2H), 2.34 (m,
3H), 2.09 (s, 3H), 1.80-2.18 (m, 8H), 1.34-1.74 (m, 7H), 1.33 (s,
3H), 1.23 (s, 3H), 1.16-1.26 (m, 2H), 1.08 (s, 3H), 1.00 (s, 6H),
0.93 (s, 3H), 0.84 (s, 3H).
Compound 21 and Compound 22:
A suspension of compound 20 (215 mg, 0.39 mmol) and Lawesson's
reagent (190 mg, 0.47 mmol) in toluene was heated at reflux for 30
min. After cooling to room temperature, the reaction mixture was
purified by column chromatography (silica gel, 0% to 65% EtOAc in
hexanes) to give product 21 (21 mg, 10% yield) as a light yellow
foam solid: m/z 550.3 (M+1). From the column, compound 22 (60 mg,
29% yield) was also obtained as a white foam solid: m/z 534.3
(M+1).
Compound 23:
NaOMe (25 w/w % solution in MeOH, 17 .mu.L, 0.074 mmol) was added
to a solution of compound 21 (33 mg, 0.060 mmol) in MeOH (0.6 mL)
at room temperature. The reaction was then heated to 55.degree. C.,
and stirred for 1 h. After cooling to 0.degree. C., t-BuOMe and 1 N
(aq.) HCl were added, and stirred for 5 min. The reaction mixture
was transferred to a separatory funnel, which was extracted with
EtOAc. The combined EtOAc extracts were washed with water, dried
over MgSO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 45% EtOAc in
hexanes) to give product 23 (24 mg, 73% yield) as a white foam
solid: m/z 550.3 (M+1). Compound 23 is an isomeric mixture of
ketone and enol forms.
Compound 63274:
To a solution of compound 23 (23 mg, 0.041 mmol) in DMF (0.3 mL)
was added 1,3-dibromo-5,5-dimethylhydantion (6.1 mg, 0.021 mmol) at
0.degree. C., and the reaction was stirred at 0.degree. C. for 1 h.
Pyridine (14 .mu.L, 0.17 mmol) was then added, and the mixture was
heated at 55.degree. C. for 3 h. After cooling to room temperature,
the reaction was diluted with EtOAc, and transferred to a
reparatory funnel, which was then washed with Na.sub.2SO.sub.3
(aq.) solution and water. The organic layer was separated, which
was dried over MgSO.sub.4, filtered, and concentrated. The crude
product was purified by column chromatography (silica gel, 0% to
45% EtOAc in hexanes) to give product 63274 (18 mg, 79% yield) as a
white foam solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.61
(s, 1H), 2.97 (d, 1H, J=4.4 Hz), 2.89 (m, 1H), 2.74 (s, 3H), 2.42
(dd, 1H, J=4.8, 16.4 Hz), 2.29 (dd, 1H, J=13.6, 16.4 Hz), 2.29 (m,
1H), 2.02 (m, 1H), 1.94 (dd, 1H, J=4.8, 13.2 Hz), 1.77-1.91 (m,
3H), 1.52-1.66 (m, 4H), 1.36-1.50 (m, 4H), 1.26 (m, 1H), 1.19 (s,
3H), 1.13 (m, 1H), 1.11 (s, 3H), 1.09 (s, 3H), 1.02 (s, 3H), 1.00
(s, 3H), 0.96 (s, 3H), 0.80 (s, 3H); m/z 548.3 (M+1).
Compound 24:
LiAlH.sub.4 solution (1.0 M in THF, 42 mL, 42 mmol) was added to a
solution of compound 1 (5.0 g, 10.3 mmol) in THF (100 mL) at room
temperature under N.sub.2. After stirring for 20 min at room
temperature, LiAlH.sub.4 solution (1.0 M in THF, 21 mL, 21 mmol)
was added again and the reaction mixture was refluxed for 1 h.
After cooling to 0.degree. C., water (10 mL) was added dropwise,
followed by the addition of 1N HCl (aq) (300 mL). The mixture was
extracted with EtOAc. The combined extracts were washed with water,
dried with MgSO.sub.4, and concentrated. The residue obtained was
mixed with CH.sub.2Cl.sub.2 (200 mL). The white solid that
precipitated was collected by filtration and washed with
CH.sub.2Cl.sub.2 (2.times.100 mL) to give compound 24 (500 mg, 10%)
as a white solid. The combined filtrate was loaded on a silica gel
column and eluted with 0% to 100% EtOAc in hexanes to give
additional compound 24 (800 mg, 17%) as a white solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 3.79 (m, 1H), 3.54 (m, 2H), 3.20 (dd,
1H, J=4.8, 10.8 Hz), 1.98 (m, 1H), 1.12-1.88 (m, 23H), 1.03 (s,
3H), 0.98 (s, 6H), 0.91 (s, 3H), 0.86 (s, 3H), 0.85 (s, 3H), 0.77
(s, 3H), 0.65-1.10 (m, 3H); m/z 443.3 (M-H.sub.2O+1), 425.3 (100%,
M-2.times.H.sub.2O+1).
Compound 25:
TEMPO (27 mg.times.4, 0.17 mmol.times.4) and IPh(OAc).sub.2 (563
mg.times.4, 1.74 mmol.times.4) were added to a white slurry of
compound 24 (725 mg, 1.59 mmol) in CH.sub.2Cl.sub.2 (200 mL) and
water (0.1 mL) at 0 h, 2 h, 24 h and 48 h at room temperature.
After stirring at room temperature for 72 h (overall reaction
time), the reaction mixture turned into a clear pink solution,
which was then transferred to a separatory funnel and washed with
Na.sub.2SO.sub.3 (aq) solution. The organic phase was separated,
dried over MgSO.sub.4, filtered, and evaporated. The residue was
purified by silica gel chromatography (0% to 75% EtOAc in hexanes)
to give compound 25 (560 mg, 77%) as a white solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 9.37 (d, 1H, J=1.2 Hz), 3.77 (m, 1H),
3.18 (dd, 1H, J=4.8, 11.2 Hz), 2.51 (m, 1H), 0.98-1.87 (m, 23H),
0.97 (s, 3H), 0.96 (s, 3H), 0.94 (s, 3H), 0.92 (m, 1H), 0.90 (s,
3H), 0.86 (s, 3H), 0.82 (s, 3H), 0.75 (s, 3H), 0.65 (m, 1H); m/z
441.3 (M-H.sub.2O+1), 423.3 (M-2.times.H.sub.2O+1).
Compound 26:
To a stirred suspension of (Ph.sub.3PCH.sub.2Cl)Cl (4.224 g, 12.1
mmol) in THF (13 mL) was added a solution of n-BuLi (4.8 mL, 11.64
mmol, 2.5 M in Hexanes) dropwise within 5 minutes at 0.degree. C.,
followed by the addition of HMPA (2.4 mL). The reaction was stirred
at r.t. for 20 minutes and then Compound 25 (1.332 g, 2.90 mmol) in
THF (13.0 mL) was added within 1 minute. The reaction mixture was
stirred at room temperature for 2 h, then quenched with HCl (1N, 20
mL) and extracted with EtOAc (100 mL). The organic phase was washed
by HCl (1N, 10 mL), NaCl (Sat., 20 mL), dried over
Na.sub.2SO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 10% to 30%
EtOAc in hexanes) to give compound 26 (1.2508 g, 87.8%, a mixture
of E/Z isomers) as a white solid:
Compound 27:
To a stirred solution of 26 (1.2508 g, 2.55 mmol) in THF (17 mL)
was added a solution of MeLi (5.16 mL, 15.44 mmol, 3 M in
CH.sub.2(OEt).sub.2) dropwise within 1 minute at 0.degree. C. The
mixture was then stirred at room temperature for 28 h and quenched
with HCl (1N, 15 mL). The aqueous solution was extracted with EtOAc
(2.times.100 mL). The combined organic phase was washed with water,
NaCl (sat.), dried over Na.sub.2SO.sub.4, filtered, and
concentrated to give compound 27 (1.0630 g, 91.7%) as a white
solid: m/z 437.3 (M-OH).
Compound 28:
To a stirred mixture of 27 (881.7 mg, 1.94 mmol), NaOAc (628.6 mg,
4 eq.) in CH.sub.2Cl.sub.2 (40 mL) was added PCC (1.257 g, 3 eq.)
in one-portion at room temperature. The mixture was then stirred at
room temperature for 5 h and diluted with a solvent mixture of
EtOAc/Hexanes (1:1, 50 mL). The mixture was directly loaded on a
silica gel pad, which was then eluted throughout with a solvent
mixture of EtOAc/Hexanes (1:1). The eluate was collected and
concentrated to give a colorless crystalline product. This crude
product was purified by column chromatography (silica gel, 0% to
10% to 25% EtOAc in hexanes) to give compound 28 (685 mg, 78.8%) as
a white solid: m/z 451.3 (M+1).
Compound 29:
To a stirred suspension of 28 (22.0 mg, 0.0488 mmol) in HCO.sub.2Et
(0.118 mL, 1.46 mmol) was added a solution of MeONa (0.167 mL,
0.732 mmol, 25% w/w in MeOH) at 0.degree. C. The mixture was then
stirred at room temperature for 25 h, diluted with TBME (1.4 mL)
and quenched with HCl (0.126 mL, conc.) followed by water (3 mL).
The aqueous solution was extracted with EtOAc (10 mL). The combined
organic phase was washed with brine (5 mL), dried over
Na.sub.2SO.sub.4, filtered, and concentrated to give compound 29 as
a light yellow foam, which was used directly in the next step.
Compound 30:
Compound 29 was dissolved in EtOH (2.1 mL). To this solution were
added NH.sub.2OH.HCl (5.1 mg, 0.0732 mmol) and H.sub.2O (0.27 mL)
at room temperature. The mixture was heated at 60.degree. C. for 18
h and then cooled to room temperature. The organic volatiles were
removed in vacuo. The remaining mixture was extracted with EtOAc
(10 mL). The organic phase was washed with water, brine, dried over
Na.sub.2SO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 10% to 25%
EtOAc in hexanes) to give 30 (20.8 mg, 89.6% from 6) as a colorless
crystalline solid: m/z 476.3 (M+1).
Compound 31:
To a stirred suspension of 30 (20.8 mg, 0.0437 mmol) in a solvent
mixture of MeOH (0.66 mL) and THF (0.11 mL) was added a solution of
MeONa (23.8 .mu.L, 0.105 mmol, 25% w/w in MeOH) at 55.degree. C.
The mixture was then stirred at 55.degree. C. for 3 h, cooled to
room temperature and quenched with 1N HCl (aq) (5 mL). The mixture
was extracted by EtOAc (15 mL). The organic phase was washed with
brine, dried over Na.sub.2SO.sub.4, filtered, and concentrated to
give compound 31 as a light yellow foam: m/z 476.3 (M-17).
Compound 63303:
Compound 31 was dissolved in benzene (2 mL). To this solution was
added a solution of DDQ (10.4 mg, 0.0458 mmol) in benzene (1 mL) at
85.degree. C. The mixture was stirred at 85.degree. C. for 1.5 h,
cooled to room temperature, and quenched with sat. NaHCO.sub.3 (aq)
(5 mL). The mixture was extracted with EtOAc (30 mL). The organic
phase was washed with sat. NaHCO.sub.3 (aq) and brine, then was
dried over Na.sub.2SO.sub.4, filtered, and concentrated to give a
solid residue (a mixture of starting material and desired product),
which was then dissolved in pyridine (0.5 mL). To this solution
were added Ac.sub.2O (50 .mu.L) and DMAP (cat.) at room
temperature. The mixture was stirred at room temperature for 30 min
and then quenched with NaHCO.sub.3 (sat.). The mixture was
extracted with EtOAc (20 mL). The organic phase was washed with
NaHCO.sub.3 (sat.), HCl (1N), brine, dried over Na.sub.2SO.sub.4,
filtered, and concentrated to give a crude mixture, which was
purified by column chromatography (silica gel, 0% to 10% to 25%
EtOAc in hexanes) to give 63303 (8.1 mg, 39.1% from 30) as a
colorless solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.66 (s,
1H), 3.25 (d, 1H, J=4.0 Hz), 2.25-2.52 (m, 3H), 2.22 (s, 1H),
1.78-2.15 (m, 5H), 1.44-1.76 (m, 9H), 1.08-1.36 (m, 2H), 1.29 (s,
3H), 1.23 (s, 3H), 1.19 (s, 3H), 1.17 (s, 3H), 0.89 (s, 3H), 0.94
(s, 3H), 0.91 (s, 3H); m/z 474.3 (M+1).
Compound 32:
Compound 31 (95 mg, 0.2 mmol) was dissolved in a solvent mixture of
acetone (3.5 mL) and water (1.5 mL). To this solution were added
HgSO.sub.4 (5.9 mg, 0.02 mmol) and H.sub.2SO.sub.4 (2 drops, conc.)
at room temperature. The mixture was stirred at 55.degree. C. for
20 h, cooled to room temperature, and quenched with water (20 mL)
and 1 N HCl (aq) (10 mL). The mixture was extracted with EtOAc (30
mL). The organic phase was washed with 1 N HCl (aq), water, sat.
NaHCO.sub.3 (aq), brine, dried over Na.sub.2SO.sub.4, filtered, and
concentrated to give a white solid, which was purified by column
chromatography (silica gel, 0% to 10% to 25% EtOAc in hexanes) to
give compound 32 (90.3 mg, 91.5%) as a white foam: m/z 494.3
(M+1).
Compound 63308:
The procedure described for the synthesis of product 63274 from
compound 23 was then employed to convert compound 32 into product
TX63308 (36.1 mg, 73.4%) as a white foam: .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.65 (s, 1H), 2.75-2.85 (m, 1H), 2.67 (d, 1H,
J=4.4 Hz), 2.44 (dt, 1H, J=16.4, 4.8 Hz), 2.35 (dt, 1H, J=16.0,
13.2 Hz), 2.16 (s, 3H), 1.92-2.06 (m, 3H), 1.30-1.76 (m, 12H),
1.18-1.29 (m, 1H), 1.22 (s, 3H), 1.16 (s, 3H), 1.15 (s, 3H), 1.04
(s, 3H), 0.97 (s, 3H), 0.96 (s, 3H), 0.93 (s, 3H); m/z 492.3
(M+1).
Compound 63323:
1,8-Diazabicyclo[5,4,0]undec-7-ene (0.18 mL, 1.204 mmol) was added
to a suspension of compound 402-51 (402 mg, 0.814 mmol) in toluene
(5.4 mL) at room temperature. After stirring for 2 min, benzyl
bromide (0.12 mL, 1.009 mmol) was added. The reaction mixture was
heated at 100.degree. C. for 6 h, after which it was cooled to room
temperature. The reaction was then diluted with EtOAc, and was
transferred to a separatory funnel, which was washed with 1 N HCl
(aq) and brine. The organic extracts were separated, which was
dried over Na.sub.2SO.sub.4, filtered and concentrated. The crude
product was purified by column chromatography (silica gel, 0% to
20% EtOAc in hexanes) to give product 63323 (308 mg, 65% yield) as
a pale yellow foam solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta.
7.56 (s, 1H), 7.30-7.37 (m, 5H), 5.21 (d, 1H, J=12.4 Hz), 5.07 (d,
1H, J=12.4 Hz), 2.84 (m, 1H), 2.47 (d, 1H, J=4.0 Hz), 2.36 (dd, 1H,
J=4.4, 16.0 Hz), 2.17 (dd, 1H, J=13.6, 16.0 Hz), 1.81-1.92 (m, 4H),
1.20-1.72 (m, 12H), 1.19 (s, 3H), 1.12 (s, 3H), 1.07 (s, 3H), 0.99
(s, 3H), 0.90 (s, 6H), 0.65 (s, 3H); m/z 584.4 (M+1).
Compound 63325:
MeONH.sub.2--HCl (109 mg, 1.305 mmol), water (0.4 mL) and Et.sub.3N
(0.24 mL, 1.722 mmol) were added sequentially to a solution of
compound 3 (439 mg, 0.857 mmol) in THF (4.2 mL) at room
temperature. The reaction was then heated at 40.degree. C. for 4 h.
After cooling to room temperature, the reaction was diluted with
EtOAc, and was transferred to a separatory funnel, which was washed
with 1 N HCl (aq) and brine. The organic extracts were separated,
which was dried over Na.sub.2SO.sub.4, filtered and concentrated.
The crude product was purified by column chromatography (silica
gel, 10% to 75% EtOAc in hexanes) to give product 63325 (165 mg,
37% yield) as a white solid: .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 8.37 (s, 1H), 7.63 (s, 1H), 3.76 (s, 3H), 2.86 (d, 1H,
J=4.0 Hz), 2.71 (m, 1H), 2.44 (dd, 1H, J=4.8, 16.4 Hz), 2.34 (dd,
1H, J=13.2, 16.4 Hz), 1.91-2.08 (m, 3H), 1.74-1.90 (m, 2H),
1.59-1.68 (m, 3H), 1.40-1.50 (m, 4H), 1.21 (s, 3H), 1.18-1.38 (m,
4H), 1.15 (s, 3H), 1.14 (s, 3H), 1.12 (s, 3H), 0.97 (s, 3H), 0.96
(s, 3H), 0.91 (s, 3H); m/z 523.4 (M+1).
Compound 63326:
Me.sub.2NH (2.0 M solution in THF, 1.23 mL, 2.460 mmol) was added
to a solution of compound 3 (408 mg, 0.797 mmol) in THF (4.1 mL) at
room temperature. The reaction was then heated at 40.degree. C. for
71 h. After cooling to room temperature, the reaction was diluted
with EtOAc, and was transferred to a separatory funnel, which was
washed with 1 N HCl (aq) and brine. The organic extracts were
separated, which was dried over Na.sub.2SO.sub.4, filtered and
concentrated. The crude product was purified by column
chromatography (silica gel, 10% to 70% EtOAc in hexanes) to give
product 63326 (254 mg, 61% yield) as a white solid: .sup.1H NMR
(400 MHz, CDCl.sub.3) .delta. 7.64 (s, 1H), 3.06 (s, 6H), 2.97 (m,
1H), 2.29-2.42 (m, 2H), 1.94-2.06 (m, 3H), 1.74-1.85 (m, 2H),
1.61-1.67 (m, 5H), 1.24-1.54 (m, 6H), 1.20 (s, 3H), 1.14 (s, 3H),
1.13 (s, 3H), 1.10 (m, 1H), 1.05 (s, 3H), 0.99 (s, 3H), 0.96 (s,
3H), 0.91 (s, 3H); m/z 521.4 (M+1).
Compound 33:
NH.sub.2OH--HCl (705 mg, 10.145 mmol), NaOAc (1.169 mg, 14.251
mmol) and water (3.3 mL) were added to a suspension of compound
402-49 (520 mg, 1.020 mmol) in EtOH (9.8 mL) at room temperature.
The reaction mixture was heated at 80.degree. C. for 27 h, after
which it was cooled to room temperature. The reaction mixture was
transferred to a separatory funnel, which was extracted with EtOAc.
The combined organic extracts were washed with water and brine,
then dried over Na.sub.2SO.sub.4, filtered and concentrated. The
crude product was purified by column chromatography (silica gel, 0%
to 20% EtOAc in hexanes) to give product 33 (387 mg, 72% yield) as
a white solid: m/z 525.3 (M+1).
Compound 34:
NaOMe (25 w/w % solution in MeOH, 0.12 mL, 0.525 mmol) was added to
a solution of compound 33 (128 mg, 0.244 mmol) in MeOH (1.2 mL) at
room temperature. The reaction was then heated to 55.degree. C. and
stirred for 1 h. After cooling to room temperature, the reaction
was diluted with t-BuOMe (3 mL) and was cooled to 0.degree. C. 1 N
HCl (aq) (5 mL) was added. After stirring for another 5 min, the
reaction mixture was transferred to a separatory funnel, which was
extracted with EtOAc. The combined EtOAc extracts were washed with
water, dried over Na.sub.2SO.sub.4, filtered, and concentrated to
give product 34 (131 mg) as a white solid: m/z 525.3 (M+1).
Compound 34 is an isomeric mixture of C3 ketone and enol forms.
Compound 63295:
1,3-Dibromo-5,5-dimethylhydantoin (40 mg, 0.140 mmol) in DMF (0.5
mL) was added to a solution of compound 34 (126 mg, 0.240 mmol) in
DMF (1.6 mL) at 0.degree. C. After stirring at 0.degree. C. for 40
min, the reaction was treated with pyridine (40 .mu.L, 0.495 mmol),
and was heated at 55.degree. C. for 7 h. After cooling to room
temperature, brine was added, and the reaction mixture was
transferred to a separatory funnel, which was extracted with EtOAc.
The combined organic extracts were washed with brine, 10%
Na.sub.2SO.sub.3 (aq) solution, 1 N HCl (aq), and water. The
organic layer was separated, which was dried over Na.sub.2SO.sub.4,
filtered, and concentrated. The crude product was purified by
column chromatography (silica gel, 0% to 20% EtOAc in hexanes) to
give product 63295 (34 mg, 27% yield from 33) as a white solid:
.sup.1H NMR (400 MHz, CDCl.sub.3) .delta. 7.83 (s, 1H), 3.68 (s,
3H), 3.36 (dd, 1H, J=16.8, 4.8 Hz), 2.82-2.91 (m, 1H), 2.54 (d, 1H,
J=3.6 Hz), 1.76-2.06 (m, 4H), 1.52-1.74 (m, 6H), 1.04-1.50 (m, 8H),
1.20 (s, 3H), 1.15 (s, 3H), 1.13 (s, 3H), 0.93 (s, 3H), 0.92 (s,
6H), 0.90 (s, 3H); m/z 523.3 (M+1);
Compound 35:
POCl.sub.3 (0.14 mL, 1.502 mmol) was added to a solution of
compound 33 (200 mg, 0.381 mmol) in pyridine (1.9 mL) at room
temperature. After stirring for 5 h, the reaction mixture was
diluted with EtOAc (5 mL), and was quenched with 1 N HCl (aq) (5
mL). The reaction mixture was transferred to a separatory funnel,
which was extracted with EtOAc. The combined organic extracts were
washed with 1 N HCl (aq) and brine, then dried over
Na.sub.2SO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 50% EtOAc in
hexanes) to give product 35 (185 mg, 75% yield) as a colorless
glassy solid: m/z 525.4 (M+1);
Compound 36:
NaOMe (25 w/w % solution in MeOH, 0.17 mL, 0.743 mmol) was added to
a solution of compound 35 (177 mg, 0.337 mmol) in MeOH (1.7 mL) at
room temperature. The reaction was then heated to 55.degree. C. and
stirred for 4 h. After cooling to 0.degree. C., t-BuOMe and 1 N HCl
(aq) were added, and the reaction mixture was stirred for 5 min.
The reaction mixture was then transferred to a separatory funnel,
which was extracted with EtOAc. The combined EtOAc extracts were
washed with 1 N HCl (aq) and brine, then dried over
Na.sub.2SO.sub.4, filtered, and concentrated to give product 36
(164 mg, 93% yield) as a white solid: m/z 525.4 (M+1). Compound 36
is an isomeric mixture of C3 ketone and enol forms.
Compound 63296:
1,3-Dibromo-5,5-dimethylhydantion (53 mg, 0.185 mmol) in DMF (0.8
mL) was added to a solution of compound 36 (163 mg, 0.311 mmol) in
DMF (2.1 mL) at 0.degree. C. After stirring at 0.degree. C. for 1
h, the reaction was treated with pyridine (50 .mu.L, 0.618 mmol)
and was heated at 55.degree. C. for 23 h. After cooling to room
temperature, brine was added, and the reaction mixture was
transferred to a separatory funnel, which was extracted with EtOAc.
The combined organic extracts were washed with brine, 10%
Na.sub.2SO.sub.3 (aq) solution, 1 N HCl (aq), and water, then dried
over Na.sub.2SO.sub.4, filtered, and concentrated to give product
63296 (150 mg, 93% yield) as a white solid: .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.88 (s, 1H), 5.57 (d, 1H, J=4.8 Hz), 4.06 (t,
1H, J=6.0 Hz), 3.73 (s, 3H), 2.68 (dd, 1H, J=14.4, 10.4 Hz),
2.48-2.60 (m, 1H), 2.13 (d, 1H, J=14.4 Hz), 1.65-1.87 (m, 3H),
1.19-1.64 (m, 13H), 1.17 (s, 3H), 1.13 (s, 3H), 1.11 (s, 3H), 1.06
(s, 3H), 1.00 (s, 3H), 0.97 (s, 3H), 0.88 (s, 3H); m/z 523.3
(M+1).
Compound 37:
m-CPBA (77%, 7.04 g, 31.52 mmol) was added to a solution of
compound 402-49 (1.60 g, 3.15 mmol) in CH.sub.2Cl.sub.2 (28 mL) at
room temperature. After stirring for 8 h, additional m-CPBA (77%,
3.52 g, 15.71 mmol) was added, and the reaction was stirred for
another 40 h. Na.sub.2SO.sub.3 (aq.) solution was then added. After
another 10 min, the reaction mixture was transferred to a
separatory funnel, which was extracted with EtOAc. The combined
EtOAc extracts were washed with NaHCO.sub.3 (aq.) solution, dried
over MgSO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 45% EtOAc in
hexanes) to give product 37 (358 mg, 22% yield) as a white foam
solid: m/z 526.3 (M+1).
Compound 38:
NaOMe (25 w/w % solution in MeOH, 20 .mu.L, 0.087 mmol) was added
to a solution of compound 37 (38 mg, 0.072 mmol) in MeOH (0.7 mL)
at room temperature. The reaction was then heated to 55.degree. C.,
and stirred for 2 h. After cooling to 0.degree. C., t-BuOMe and 1 N
(aq.) HCl were added. The reaction mixture was then transferred to
a separatory funnel, which was extracted with EtOAc. The combined
EtOAc extracts were washed with water, dried over MgSO.sub.4,
filtered, and concentrated. The crude product was purified by
column chromatography (silica gel, 0% to 50% EtOAc in hexanes) to
give product 38 (25 mg, 66% yield) as a white foam solid: m/z 526.4
(M+1). Compound 38 is an isomeric mixture of C3 ketone and enol
forms.
Compound 63263:
A solution of 1,3-dibromo-5,5-dimethylhydantion (6.9 mg, 0.024
mmol) in DMF (0.2 mL) was added to a solution of compound 38 (25
mg, 0.048 mmol) in DMF (0.8 mL) at 0.degree. C. After stirring at
0.degree. C. for 1 h, the reaction was treated with pyridine (12
.mu.L, 0.15 mmol), and was heated at 55.degree. C. for 3 h. After
cooling to room temperature, the reaction was diluted with EtOAc,
and transferred to a separatory funnel, which was then washed with
Na.sub.2SO.sub.3 (aq.) solution, 1N (aq.) HCl and water. The
organic extract was separated, dried over MgSO.sub.4, filtered, and
concentrated. The crude product was purified by column
chromatography (silica gel, 0% to 45% EtOAc in hexanes) to give
product 63263 (18 mg) as a white foam solid: .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.83 (s, 1H), 4.95 (d, 1H, J=7.2 Hz), 3.73 (s,
3H), 2.79 (m, 2H), 2.37 (d, 1H, J=14.4 Hz), 1.92 (m, 2H), 1.77 (d,
1H, J=10.4 Hz), 1.44-1.74 (m, 8H), 1.20-1.41 (m, 5H), 1.19 (s, 3H),
1.15 (s, 3H), 1.13 (s, 3H), 1.08 (s, 3H), 1.07 (s, 3H), 0.96 (s,
3H), 0.89 (s, 3H); m/z 524.3 (M+1).
Compound 39:
LiAlH.sub.4 (2.0 M in THF, 48 .mu.L, 0.096 mmol) was added to a
solution of compound 37 (50 mg, 0.095 mmol) in THF (0.95 mL) at
0.degree. C. After stirring for 40 min, the reaction was quenched
by adding water (1 mL) carefully. After stirring at room
temperature for 10 min, the reaction mixture was transferred to a
separatory funnel, which was extracted with EtOAc. The combined
organic extracts were washed with 1N (aq.) HCl, and water, dried
over MgSO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 40% EtOAc in
hexanes) to give product 39 (31 mg, 62% yield) as a white foam
solid: m/z 528.3 (M+1). The stereochemical configuration of C12 was
not assigned.
Compound 40:
NaOMe (25 w/w % solution in MeOH, 16 .mu.L, 0.070 mmol) was added
to a solution of compound 39 (30 mg, 0.057 mmol) in MeOH (0.6 mL)
at room temperature. The reaction was then heated to 55.degree. C.,
and stirred for 2 h. After cooling to 0.degree. C., t-BuOMe and 1 N
(aq.) HCl were added. The reaction mixture was then transferred to
a separatory funnel, which was extracted with EtOAc. The combined
EtOAc extracts were washed with water, dried over MgSO.sub.4,
filtered, and concentrated. The crude product was purified by
column chromatography (silica gel, 0% to 40% EtOAc in hexanes) to
give product 40 (25 mg, 83% yield) as a white foam solid: m/z 510.3
(M-18+1). Compound 40 is an isomeric mixture of C3 ketone and enol
forms. The stereochemical configuration of C12 was not
assigned.
Compound 63289:
1,3-Dibromo-5,5-dimethylhydantion (6.8 mg, 0.024 mmol) was added to
a solution of compound 40 (25 mg, 0.047 mmol) in DMF (0.47 mL) at
0.degree. C. After stirring at 0.degree. C. for 1 h, the reaction
was treated with pyridine (12 .mu.L, 0.15 mmol), and was heated at
55.degree. C. for 3 h. After cooling to room temperature, the
reaction was diluted with EtOAc, and transferred to a separatory
funnel, which was then washed with Na.sub.2SO.sub.3 (aq.) solution,
1N (aq.) HCl and water. The organic extract was separated, dried
over MgSO.sub.4, filtered, and concentrated. The crude product was
purified by column chromatography (silica gel, 0% to 40% EtOAc in
hexanes) to give partially purified product 63289 (16 mg), which
was purified again by preparative TLC plate (silica gel, eluted
with 8% EtOAc in hexanes) to give product 63289 (10 mg, 40% yield)
as a white foam solid: .sup.1H NMR (400 MHz, CDCl.sub.3) .delta.
7.64 (s, 1H), 5.12 (m, 1H), 4.33 (d, 1H, J=6.8 Hz), 3.71 (s, 3H),
2.54 (m, 1H), 2.43 (d, 1H, J=2.8 Hz), 1.21-1.96 (m, 18H), 1.19 (s,
6H), 1.13 (s, 3H), 1.05 (s, 3H), 1.02 (s, 3H), 0.95 (s, 3H), 0.88
(s, 3H); m/z 508.3 (M-18+1). The stereochemical configuration of
C12 was not assigned.
Example 4
Uptake of 404-02 into CNS and Lungs of Monkeys
Plasma Concentrations Following Oral Dosing:
Compound 404-02 shows high uptake in the CNS and lung in monkeys
following oral dosing: 2 male and 2 female cynomolgus monkeys were
administered 404-02 at 0.5, 5, 25 or 75 mg/kg/day doses via oral
gavage. Doses were prepared in sesame oil and individualized to
body weight on the day of dosing. Blood was drawn prior to dosing
and at 1, 2, 4, 8 and 24 hours post-dose on days 1 and 12. Blood
samples were collected from the femoral artery/vein for
determination of 404-02 plasma concentrations. Blood was placed in
tubes containing K3EDTA and stored on ice until centrifugation at
room temperature. The isolated plasma was transferred to cryovials
and stored at -80.degree. C. until sample processing and LC-MS/MS
analysis. Extracted plasma standard curves were prepared from fresh
stock solutions and analyzed prior to study samples. Summary
results are shown in Tables 2a and 2b.
Population mean pharmacokinetic parameter estimates were obtained
by performing non-compartmental analysis of the 404-02 plasma
concentration-versus-time data using WinNonlin.TM. software version
5.2. Across the investigated dose range, 404-02 demonstrated
dose-dependent kinetics with increased oral clearance (Cl/F),
reciprocal shortening of elimination half-life (T.sub.1/2) and an
increasing apparent volume of distribution (V.sub.z/F) with
increasing dose. A 1.6-fold increase in the area under the
concentration versus time curve over 24 hours (AUC.sub.0-24 hr) was
observed at the 75 mg/kg dose level after 12 days of dosing
compared to the corresponding AUC on day 1. Accumulation was not
observed at any of the other dose levels. The observed mean maximum
plasma concentration (C.sub.max) for the 0.5, 5, 25 and 75
mg/kg/day dose groups on day 12 were 4.6, 12.7, 17.5 and 48.6 nM
404-02, respectively.
Table 2a shows the population mean plasma pharmacokinetics of
404-02 in cynomolgus monkey on day 1 of study (n=4).
Pharmacokinetic parameters obtained using non-compartmental
analysis, WinNonlin.TM. version 5.2.
TABLE-US-00010 TABLE 2a Day 1 Plasma Pharmacokinetics of 404-02:
Mean Maximum 404-02 Mean Maximum 404-02 Dose Plasma Concentration
Plasma Concentration (mg/kg/d) (ng/mL) .+-. SEM (nM) .+-. SEM 0.5
1.95 .+-. 0.35 3.4 .+-. 0.61 5 10.1 .+-. 3.8 17.6 .+-. 6.6 25 16.8
.+-. 1.3 29.3 .+-. 2.3 75 19.4 .+-. 2.7 33.8 .+-. 4.7
Table 2b shows the population mean plasma pharmacokinetics of
404-02 in cynomolgus monkey on day 12 of study (n=4).
Pharmacokinetic parameters were obtained using non-compartmental
analysis using WinNonlin.TM. version 5.2 software.
TABLE-US-00011 TABLE 2b Day 12 Plasma Pharmacokinetics of 404-02:
Mean Maximum 404-02 Mean Maximum 404-02 Dose Plasma Concentration
Plasma Concentration (mg/kg/d) (ng/mL) .+-. SEM (nM) .+-. SEM 0.5
2.65 .+-. 0.6 4.6 .+-. 1.0 5 7.3 .+-. 1.7 12.7 .+-. 2.9 25 10.0
.+-. 3.7 17.5 .+-. 6.4 75 28.0 .+-. 12.2 48.6 .+-. 12.2
CNS and Lung Concentrations Following Oral Dosing:
2 male and 2 female cynomolgus monkeys were administered 404-02 at
0.5, 5, 25 or 75 mg/kg/day doses via oral gavage, in addition to a
control (non-treatment) group with 2 animals per sex. Animals were
sacrificed approximately 3 hours after dosing on day 15 following
harvest of brain and lung tissues. Each sample collected was rinsed
in 1.times. isotonic phosphate buffered saline and blotted dry
before weighing. Harvested tissue slices were transferred to
cryovials and stored at -80.degree. C. until processing and
LC-MS/MS analysis. Standard curves were derived for 404-02 in
homogenates of these tissues and were used to quantify the day 15
samples.
The concentration of 404-02 necessary for 50% suppression of nitric
oxide (NO) production in macrophages stimulated with
interferon-gamma is approximately 45 nM. As evidenced by the data,
0.5 mg/kg administered orally as the lowest dose in this study for
15 days resulted in a mean 404-02 CNS concentration of 2,162 nM,
which markedly exceeds the IC.sub.50 value for NO production in
vitro. The CNS penetration provides a large therapeutic margin at
all doses tested (Table 2c). For instance, the mean 404-02 CNS
concentrations following administration of 0.5, 5, 25 and 75
mg/kg/day 404-02 indicates a 48-, 44-, 37- and 75-fold excess
compared to the dose necessary to suppress inflammation in vitro.
For comparison, the mean 404-02 lung tissue exposure at 75
mg/kg/day 404-02 showed a 133-fold increase (Table 2d). Non-linear
disposition of 404-02 in CNS and lung tissue was observed
suggesting 404-02 is taken up by a saturable mechanism(s) for
transport across membranes and/or intracellular binding. In
addition, concentrations of 404-02 in monkey CNS and lung tissue
exceed plasma levels. Table 2c shows the population mean 404-02
cynomolgus monkey CNS tissue exposure on day 15. Table 2d shows the
population mean 404-02 cynomolgus monkey lung tissue content on day
15.
TABLE-US-00012 TABLE 2c Day 15 CNS Tissue Content/Concentration of
404-02: Mean 404-02 CNS Mean 404-02 CNS Dose Tissue Content Tissue
Concentration (mg/kg/day) (ng/g) .+-. SEM (nM)* .+-. SEM 0.5 1198
.+-. 712 2087 .+-. 1239 5 1090 .+-. 564 1900 .+-. 982 25 930 .+-.
352 1618 .+-. 612 75 1898 .+-. 496 3305 .+-. 864 *Conversion based
on the assumption the density of tissue is equal to water, 1
g/mL.
TABLE-US-00013 TABLE 2d Day 15 Lung Tissue Content/Concentration of
404-02: Mean 404-02 Lung Mean 404-02 Lung Dose Level Tissue Content
Tissue Concentration (mg/kg/day) (ng/g) .+-. SEM (nM)* .+-. SEM 0.5
705 .+-. 292 1227 .+-. 508 5 24 .+-. 15 55 .+-. 27 25 141 .+-. 50
246 .+-. 87 75 3405 .+-. 824 5929 .+-. 1435
Example 5
Uptake of 404-02 into CNS and Lungs of Rats
Compound 404-02 reaches high concentrations in the lung and CNS of
rats following oral dosing: To assess basic pharmacokinetic
parameters following oral dosing, 9 male and 9 female Sprague
Dawley (SD) rats were administered 404-02 at 1, 10 or 50 mg/kg
doses via oral gavage. Doses were prepared in sesame oil and
individualized to body weight on the day of dosing. Blood was drawn
at 0, 1, 2, 4, 8 and 24 hours post-dose on days 1 and 15. Blood was
collected from the orbital sinus after carbon dioxide/oxygen
inhalation for determination of 404-02 plasma concentrations.
Plasma was transferred to cryovials and stored at -80.degree. C.
until processing and LC-MS/MS analysis. Summary results for Day 1
and Day 15 are shown in Tables 2a and 2b, respectively. A standard
curve was derived for 404-02 in rat plasma, and quantification of
experimental results was based on this standard curve.
Table 3a shows population mean plasma pharmacokinetics of 404-02 in
SD rats on day 1 of study (n=9/sex/dose level). Pharmacokinetic
parameters were obtained using non-compartmental analysis,
WinNonlin.TM. version 5.2.
TABLE-US-00014 TABLE 3a Day 1 Pharmacokinetic Parameters Mean
Maximum 404-02 Mean Maximum 404-02 Dose Plasma Concentration Plasma
Concentration (mg/kg/day) (ng/mL) .+-. SEM (nM) .+-. SEM 1 3.3 .+-.
0.65 5.7 .+-. 1.1 10 66 .+-. 16.5 116 .+-. 28.7 50 713 .+-. 98.2
1241 .+-. 171
Table 3b shows the population mean plasma pharmacokinetics of
404-02 in SD rats on day 15 of study (n=9/sex/dose level).
Pharmacokinetic parameters were obtained using non-compartmental
analysis, WinNonlin.TM. version 5.2.
TABLE-US-00015 TABLE 3b Day 15 Pharmacokinetic Parameters: Mean
Maximum 404-02 Mean Maximum 404-02 Dose Plasma Concentration Plasma
Concentration (mg/kg/day) (ng/mL) .+-. SEM (nM) .+-. SEM 1 6.3 .+-.
0.9 11.0 .+-. 1.6 10 144 .+-. 24.3 .sup. 210 .+-. 42.3 50 1129 .+-.
114 1966 .+-. 199
To examine tissue concentrations following oral dosing, 5 male and
5 female Sprague Dawley (SD) rats were administered 404-02 at 1,
10, 50 or 150 mg/kg/day via oral gavage. Doses were prepared in
sesame oil and individualized to body weight on the day of dosing.
Animals were sacrificed 3 hours post-dose on day 15 of study
following harvest of brain and lung specimens. Each sample
collected was rinsed in 1.times. isotonic phosphate buffered saline
and blotted dry before weighing. Tissue slices were transferred to
cryovials and stored at -80.degree. C. until processing and
LC-MS/MS analysis. Summary results for CNS and lung samples are
shown in Tables 3c and 3d, respectively. Standard curves were
derived for 404-02 in these tissues.
TABLE-US-00016 TABLE 3c Population Mean 404-02 CNS Tissue Content
in SD Rats on Day 15. Mean 404-02 CNS Mean 404-02 CNS Dose Tissue
Content Tissue Concentration (mg/kg/day) (ng/g = ng/mL)* .+-. SEM
(nM) .+-. SEM 1 29 .+-. 6.8 51 .+-. 11.8 10 421 .+-. 99 733 .+-.
173 50 750 .+-. 108 1306 .+-. 188 150 640 .+-. 82 1114 .+-. 143
*Conversion based on the assumption the density of tissue is equal
to water, 1 g/mL.
TABLE-US-00017 TABLE 3d Population Mean 404-02 Lung Tissue Content
in SD Rats on Day 15. Mean dh404 Lung Mean dh404 Lung Dose Tissue
Content Tissue Concentration (mg/kg/day) (ng/g = ng/mL)* .+-. SEM
(nM) .+-. SEM 1 558 .+-. 123 972 .+-. 215 10 4032 .+-. 928 7020
.+-. 1616 50 7165 .+-. 1221 12474 .+-. 2126 150 9719 .+-. 1643
16921 .+-. 2860 *Conversion based on the assumption the density of
tissue is equal to water, 1 g/mL.
Example 6
Rodent Toxicity Comparison Between 402 and 402-02
A study was performed in Sprague Dawley rats using both 402 and
402-02. Animals were dosed orally once daily for 7 days. The low
dose 402 group had elevated total bilirubin and GGT levels as well
as suppressed weight gain. The high dose animals that were treated
with 402 were all sacrificed in extremis on Day 6 before study
completion. GGT and total bilirubin levels were elevated in these
animals as well. However, no toxicity as assessed by clinical
observations, weight gain, GGT, and total bilirubin was observed in
any animal treated with 402-02 (Table 4). In a second study
involving oral administration to Sprague-Dawley rats for 14 days,
402-02, achieved comparable blood levels to that of 402. However,
no significant toxicity was observed as assessed by weight loss,
clinical observations, and GGT and total bilirubin elevations
relative to controls at doses up to 1,500 mg/m.sup.2/day for 14
days, which is 50-fold higher than the MTD of RTA 402 in this
species (Table 2).
TABLE-US-00018 TABLE 4 Comparing Compounds 402-02 and 402 for
Rodent Toxicity Total Weight Bili- vs GGT rubin Timepoint/Dose
Level Survival Control (U/L) (mg/dL) 7 Day Study - Crystalline
Forms of 402 and 402-02 Vehicle Control 4/4 (100%) 100% <5 0.13
402 - 60 mg/m.sup.2/day 4/4 (100%) 35% 8.7 0.25 402 - 180
mg/m.sup.2/day 0/4 (0%) NA 5.0 0.78 402-02 - 60 mg/m.sup.2/day 4/4
(100%) 106% <5 0.20 402-02 - 180 mg/m.sup.2/day 4/4 (100%) 132%
<5 0.20 14 Day Study - Crystalline Form of 402-02 Vehicle
Control 10/10 (100%) 100% <5 0.20 402-02 - 60 mg/m.sup.2/day
10/10 (100%) 95% <5 0.24 402-02 - 180 mg/m.sup.2/day 5/5 (100%)
102% <5 0.22 402-02 - 600 mg/m.sup.2/day 5/5 (100%) 96% <5
0.20 402-02 - 1,500 mg/m.sup.2/day 5/5 (100%) 105% <5 0.20
Example 7
Toxicity Comparison in Mice
In this study, six compounds (401, 402, 404, 401-2, 402-2, and
404-2) were assessed for toxicity in mice in a 14-day study. Each
compound was formulated in sesame oil and administered daily by
oral gavage at doses of 10, 50, 100, or 250 mg/kg (n=4 per group).
At higher doses (above 10 mg/kg/day) both 401 and 402 caused at
least 50% mortality; 404 was non-toxic. In contrast, no mortality
was observed in the 402-2 and 404-2 groups and only the highest
dose of 401-02 caused any lethality (Table 5). Body weight
measurements (FIGS. 29-31) were consistent with the mortality
observations. The two highest doses of 401 and 402 were lethal
within 4 days, in contrast to the effects of 401-2 and 402-2.
TABLE-US-00019 TABLE 5 Mortality Observations in 14-Day Toxicity
Study Number Dose of Group Compound (mg/kg) Schedule N Deaths
Comments 1 vehicle QD .times. 14, 4 0 D 1-14 2 401 10 QD .times.
14, 4 0 D 1-14 3 401 50 QD .times. 14, 4 2 D 1-14 4 401 100 QD
.times. 14, 4 4 D 1-14 5 401 250 QD .times. 14, 4 4 D 1-14 6 401-02
10 QD .times. 14, 4 0 D 1-14 7 401-02 50 QD .times. 14, 4 1* *Due
to D 1-14 gavage injury 8 401-02 100 QD .times. 14, 4 0 D 1-14 9
401-02 250 QD .times. 14, 4 1 Sacrificed D 1-14 due to weightless
on Day 11 10 402 10 QD .times. 14, 4 0 D 1-14 11 402 50 QD .times.
14, 4 4 D 1-14 12 402 100 QD .times. 14, 4 4 D 1-14 13 402 250 QD
.times. 14, 4 4 D 1-14 14 402-02 10 QD .times. 14, 4 0 D 1-14 15
402-02 50 QD .times. 14, 4 0 ID 1-14 15 402-02 100 QD .times. 14, 4
0 D 1-14 17 402-02 250 QD .times. 14, 4 0 D 1-14 .sup. 1a 404 10 QD
.times. 14, 4 0 D 1-14 19 404 50 QD .times. 14, 4 0 D 1-14 20 404
100 QD .times. 14, 4 0 D 1-14 21 404 250 QD .times. 14, 4 0 D 1-14
22 404-02 10 QD .times. 14, 4 0 D 1-14 23 404-02 50 QD .times. 14,
4 0 D 1-14 22 404-02 100 QD .times. 14, 4 0 D 1-14 23 404-02 250 QD
.times. 14, 4 0 D 1-14
In a second experiment, six additional compound differing only in
the saturation or non-saturation of the C ring were tested for
toxicity in mice by daily oral administration for 9 days, using
sesame oil as the vehicle. In this study, no significant toxicity
was observed. The deaths of two animals were attributed to gavage
errors during the administration of the test article. No
significant differences in weight were observed in any group
compared to the vehicle-treated controls. Results are summarized in
Table 6 below. As with compounds 402-2, 401-2 and 404-2 above,
compounds with saturation in the C ring consistently show low
toxicity in rodents. Compounds lacking saturation in the C ring
show significant rodent toxicity in some cases (e.g., 401 and 402).
Predictably low rodent toxicity provides an advantage since high
rodent toxicity can be a significant complication in conducting
preclinical studies required for development and registration of
therapeutic compounds for use in humans or non-human animals.
TABLE-US-00020 TABLE 6 Further Mouse Toxicity Results. Compound
Dose (per day, p.o.) Mortality 63112 3 mg/kg 0/5 10 mg/kg 0/5 30
mg/kg 1/5 63323 3 mg/kg 0/5 10 mg/kg 0/5 30 mg/kg 0/5 63324 3 mg/kg
0/5 10 mg/kg 0/5 30 mg/kg 0/5 63325 3 mg/kg 0/5 10 mg/kg 0/5 30
mg/kg 0/5 63166 3 mg/kg 0/5 10 mg/kg 0/5 30 mg/kg 0/5 63326 3 mg/kg
0/5 10 mg/kg 1/5 30 mg/kg 0/5
Example 8
Aqueous Solubility of Oleanolic Acid Derivatives
The aqueous solubility of the compounds shown here was determined
using the procedures outlined in Example 1.
TABLE-US-00021 Aqueous Solubility Compound ID(s) Structure (.mu.M)
63097 (402) ##STR00076## 1.46 63102 (dh404) ##STR00077## 0.06 63198
##STR00078## 163.6 63202 ##STR00079## 1.89 63208 ##STR00080## 9.49
63214 ##STR00081## 112.2 63219 ##STR00082## 13.58 63221
##STR00083## 8.78 63226 ##STR00084## 0.71 63231 ##STR00085## 1.23
63232 ##STR00086## 0.75 63237 ##STR00087## 5.16
All of the methods disclosed and claimed herein can be made and
executed without undue experimentation in light of the present
disclosure. While the compositions and methods of this invention
have been described in terms of preferred embodiments, it will be
apparent to those of skill in the art that variations may be
applied to the methods and in the steps or in the sequence of steps
of the method described herein without departing from the concept,
spirit and scope of the invention. More specifically, it will be
apparent that certain agents which are both chemically and
physiologically related may be substituted for the agents described
herein while the same or similar results would be achieved. All
such similar substitutes and modifications apparent to those
skilled in the art are deemed to be within the spirit, scope and
concept of the invention as defined by the appended claims.
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SEQUENCE LISTINGS
1
10120DNAArtificial SequenceSynthetic primer 1tccgatgggt ccttacactc
20220DNAArtificial SequenceArtificial primer 2taggctcctt cctcctttcc
20320DNAArtificial SequenceArtificial primer 3gcagcactga gtggtcaaaa
20420DNAArtificial SequenceArtificial primer 4ggtcaactgc ctcaattgct
20520DNAArtificial SequenceArtificial primer 5gctgtggcta ctgcggtatt
20620DNAArtificial SequenceArtificial primer 6atctgcctca atgacaccat
20720DNAArtificial SequenceArtificial primer 7atgagcaggt gaaagccatc
20820DNAArtificial SequenceArtificial primer 8taaaggaaac cccaacatgc
20920DNAArtificial SequenceArtificial primer 9gattacatcc tgggcctgaa
201020DNAArtificial SequenceSynthetic primer 10gagcgcagag
agaagtcgat 20
* * * * *
References